WorldWideScience

Sample records for rapid identification scheme

  1. An MLSA-based online scheme for the rapid identification of Stenotrophomonas isolates

    Directory of Open Access Journals (Sweden)

    Patrícia Locosque Ramos

    2011-06-01

    Full Text Available An online scheme to assign Stenotrophomonas isolates to genomic groups was developed using the multilocus sequence analysis (MLSA, which is based on the DNA sequencing of selected fragments of the housekeeping genes ATP synthase alpha subunit (atpA, the recombination repair protein (recA, the RNA polymerase alpha subunit (rpoA and the excision repair beta subunit (uvrB. This MLSA-based scheme was validated using eight of the 10 Stenotrophomonas species that have been previously described. The environmental and nosocomial Stenotrophomonas strains were characterised using MLSA, 16S rRNA sequencing and DNA-DNA hybridisation (DDH analyses. Strains of the same species were found to have greater than 95% concatenated sequence similarity and specific strains formed cohesive readily recognisable phylogenetic groups. Therefore, MLSA appeared to be an effective alternative methodology to amplified fragment length polymorphism fingerprint and DDH techniques. Strains of Stenotrophomonas can be readily assigned through the open database resource that was developed in the current study (www.steno.lncc.br/.

  2. Rapid Calculation of EMI Responses of Metallic Objects and Implementation in Inversion Schemes

    National Research Council Canada - National Science Library

    Paulsen, Keith

    2003-01-01

    .... Nonetheless, signal inversion schemes are impeded by a lack of rapid means for calculating the responses of possible target types in a great variety of depths and dispositions relative to the sensor...

  3. A flexible and rapid frequency selective scheme for SRS microscopy

    Science.gov (United States)

    Li, Jingting; Yue, Yuankai; Shih, Wei-Chuan

    2017-02-01

    Stimulated Raman scattering (SRS) is a label-free imaging technique suitable for studying biological systems. Due to stimulated nature by ultrafast laser pulses, SRS microscopy has the advantage of significantly higher sensitivity but often reduced spectroscopic information. In this paper, we present a newly constructed femtosecond SRS microscope with a high-speed dynamic micromirror device based pulse shaper to achieve flexible and rapid frequency selection within the C-H stretch region near 2800 to 3100 cm-1 with spectral width of 30 cm-1. This technique is applicable to lipid profiling such as cell activity mapping, lipid distribution mapping and distinction among subclasses.

  4. A color and texture based multi-level fusion scheme for ethnicity identification

    Science.gov (United States)

    Du, Hongbo; Salah, Sheerko Hma; Ahmed, Hawkar O.

    2014-05-01

    Ethnicity identification of face images is of interest in many areas of application. Different from face recognition of individuals, ethnicity identification classifies faces according to the common features of a specific ethnic group. This paper presents a multi-level fusion scheme for ethnicity identification that combines texture features of local areas of a face using local binary patterns with color features using HSV binning. The scheme fuses the decisions from a k-nearest neighbor classifier and a support vector machine classifier into a final identification decision. We have tested the scheme on a collection of face images from a number of publicly available databases. The results demonstrate the effectiveness of the combined features and improvements on accuracy of identification by the fusion scheme over the identification using individual features and other state-of-art techniques.

  5. An Extended Multilocus Sequence Typing (MLST Scheme for Rapid Direct Typing of Leptospira from Clinical Samples.

    Directory of Open Access Journals (Sweden)

    Sabrina Weiss

    2016-09-01

    Full Text Available Rapid typing of Leptospira is currently impaired by requiring time consuming culture of leptospires. The objective of this study was to develop an assay that provides multilocus sequence typing (MLST data direct from patient specimens while minimising costs for subsequent sequencing.An existing PCR based MLST scheme was modified by designing nested primers including anchors for facilitated subsequent sequencing. The assay was applied to various specimen types from patients diagnosed with leptospirosis between 2014 and 2015 in the United Kingdom (UK and the Lao Peoples Democratic Republic (Lao PDR. Of 44 clinical samples (23 serum, 6 whole blood, 3 buffy coat, 12 urine PCR positive for pathogenic Leptospira spp. at least one allele was amplified in 22 samples (50% and used for phylogenetic inference. Full allelic profiles were obtained from ten specimens, representing all sample types (23%. No nonspecific amplicons were observed in any of the samples. Of twelve PCR positive urine specimens three gave full allelic profiles (25% and two a partial profile. Phylogenetic analysis allowed for species assignment. The predominant species detected was L. interrogans (10/14 and 7/8 from UK and Lao PDR, respectively. All other species were detected in samples from only one country (Lao PDR: L. borgpetersenii [1/8]; UK: L. kirschneri [1/14], L. santarosai [1/14], L. weilii [2/14].Typing information of pathogenic Leptospira spp. was obtained directly from a variety of clinical samples using a modified MLST assay. This assay negates the need for time-consuming culture of Leptospira prior to typing and will be of use both in surveillance, as single alleles enable species determination, and outbreaks for the rapid identification of clusters.

  6. [Rapid identification system for seedlings of medicinal Chrysanthemum morifolium].

    Science.gov (United States)

    Mao, Pengfei; Guo, Qiaosheng; Wang, Tao; Shao, Qingsong

    2012-04-01

    To achieve the rapid identification for seedlings of medicinal Chrysanthemum morifolium, the discriminant equation was established and the software for rapid identification was designed. Leaf structure of medicinal Chrysanthemum of 12 cultivars was analyzed to establish the discriminant equation based on variance analysis and discriminant analysis. On this basis, the identification program and software (based on the python language) were designed. Through the analysis of variance and multiple comparisons for the 11 leaf parameter index data of 12 different cultivars, it was found that that the leaf parameters were significant different from each other and reached significant levels. The discriminant equation and the rapid identification software were set up based on the analysis of various indicators. The rapid identification system of seedlings of medicinal Chrysanthemum could be achieved through the establishment of discriminant equation combined with computer technology.

  7. Numerical simulation of the rapid intensification of Hurricane Katrina (2005): Sensitivity to boundary layer parameterization schemes

    Science.gov (United States)

    Liu, Jianjun; Zhang, Feimin; Pu, Zhaoxia

    2017-04-01

    Accurate forecasting of the intensity changes of hurricanes is an important yet challenging problem in numerical weather prediction. The rapid intensification of Hurricane Katrina (2005) before its landfall in the southern US is studied with the Advanced Research version of the WRF (Weather Research and Forecasting) model. The sensitivity of numerical simulations to two popular planetary boundary layer (PBL) schemes, the Mellor-Yamada-Janjic (MYJ) and the Yonsei University (YSU) schemes, is investigated. It is found that, compared with the YSU simulation, the simulation with the MYJ scheme produces better track and intensity evolution, better vortex structure, and more accurate landfall time and location. Large discrepancies (e.g., over 10 hPa in simulated minimum sea level pressure) are found between the two simulations during the rapid intensification period. Further diagnosis indicates that stronger surface fluxes and vertical mixing in the PBL from the simulation with the MYJ scheme lead to enhanced air-sea interaction, which helps generate more realistic simulations of the rapid intensification process. Overall, the results from this study suggest that improved representation of surface fluxes and vertical mixing in the PBL is essential for accurate prediction of hurricane intensity changes.

  8. Rapid Immunization Scheme for Spouses of Individuals Estabilished as Hepatitis B Carriers during Premarital Tests

    Directory of Open Access Journals (Sweden)

    Selma Tosun

    2012-01-01

    Full Text Available Background. The aim of this study was to monitor the cases identified as hepatitis B carriers during premarital tests, to vaccinate their prospective spouses with a rapid vaccination scheme, and to compare the anti-HBs responses with the traditional vaccination scheme. Methods. Blood samples of 1250 couple spouses were tested for HBsAg and anti-HBs. HBsAg positive cases' fiancées which were found HBV negative were administered a rapid three-dose vaccination scheme on days 0, 7, and 21. Forty controls with similar age and gender were also were administered three doses of the same vaccine. Results. Out of 1250 cases (625 couples, 46 (3.6% were HBsAg positive, and 40 of them aged between 18 and 39 were admitted to the rapid vaccination program. Conclusion. Upon determination of HBsAg positivity in premarital tests, a rapid vaccination program provides early protection, but the 6th and 12th month vaccinations are also required. Anti-HBs response should be monitored.

  9. Rapid Identification of Sporothrix Species by T3B Fingerprinting

    Science.gov (United States)

    de Oliveira, Manoel Marques Evangelista; Sampaio, Paula; Almeida-Paes, Rodrigo; Pais, Célia; Gutierrez-Galhardo, Maria Clara

    2012-01-01

    This article describes PCR fingerprinting using the universal primer T3B to distinguish among species of the Sporothrix complex, S. brasiliensis, S. globosa, S. mexicana, and S. schenckii. This methodology generated distinct banding patterns, allowing the correct identification of all 35 clinical isolates at the species level, confirmed by partial calmodulin (CAL) gene sequence analyses. This methodology is simple, reliable, rapid, and cheap, making it an ideal routine identification system for clinical mycology laboratories. PMID:22403427

  10. Rapid identification of DNA-binding proteins by mass spectrometry

    DEFF Research Database (Denmark)

    Nordhoff, E; Krogsdam, A M; Jorgensen, H F

    1999-01-01

    We report a protocol for the rapid identification of DNA-binding proteins. Immobilized DNA probes harboring a specific sequence motif are incubated with cell or nuclear extract. Proteins are analyzed directly off the solid support by matrix-assisted laser desorption/ionization time-of-flight mass...... was validated by the identification of known prokaryotic and eukaryotic DNA-binding proteins, and its use provided evidence that poly(ADP-ribose) polymerase exhibits DNA sequence-specific binding to DNA....

  11. Rapid identification of the medicinal plant Taraxacum formosanum ...

    African Journals Online (AJOL)

    Rapid identification of the medicinal plant Taraxacum formosanum and distinguishing of this plant from its adulterants by ribosomal DNA internal transcribed spacer (ITS) based DNA barcode. YC Chiang, WT Chang, MD Chen, GH Lai, HJ Chen, J Chao, MK Lin, YS Chang, YM Chou, MS Lee, MS Lee ...

  12. Two Unusual Occurrences of Trichomoniasis: Rapid Species Identification by PCR▿

    Science.gov (United States)

    Bellanger, A. P.; Cabaret, O.; Costa, J. M.; Foulet, F.; Bretagne, S.; Botterel, F.

    2008-01-01

    PCR analysis in two unusual occurrences of trichomoniasis, trichomonal empyema due to Trichomonas tenax and Trichomonas vaginalis in an infant urine sample, allowed us to obtain rapid and accurate trichomonad species identification. The weak sensitivity of wet preparations and the low viability of the flagellates can be remedied by the PCR method. PMID:18632901

  13. Development of rapid phenotypic system for the identification of ...

    Indian Academy of Sciences (India)

    Rapid and accurate identification of bacterial pathogens is a fundamental goal of clinical microbiology. The diagnosis and surveillance of diseases is dependent, to a great extent, on laboratory services, which cannot function without effective reliable reagents and diagnostics. Despite the advancement in microbiology ...

  14. Rapid identification of Candida species by confocal Raman microspectroscopy

    NARCIS (Netherlands)

    K. Maquelin (Kees); L.P. Choo-Smith; H.P. Endtz (Hubert); H.A. Bruining (Hajo); G.J. Puppels (Gerwin)

    2002-01-01

    textabstractCandida species are important nosocomial pathogens associated with high mortality rates. Rapid detection and identification of Candida species can guide a clinician at an early stage to prescribe antifungal drugs or to adjust empirical therapy when resistant species are

  15. Rapid species specific identification and subtyping of Yersinia enterocolitica by MALDI-TOF mass spectrometry.

    Science.gov (United States)

    Stephan, Roger; Cernela, Nicole; Ziegler, Dominik; Pflüger, Valentin; Tonolla, Mauro; Ravasi, Damiana; Fredriksson-Ahomaa, Maria; Hächler, Herbert

    2011-11-01

    Yersinia enterocolitica are Gram-negative pathogens and known as important causes of foodborne infections. Rapid and reliable identification of strains of the species Y. enterocolitica within the genus Yersinia and the differentiation of the pathogenic from the non-pathogenic biotypes has become increasingly important. We evaluated here the application of matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS) for rapid species identification and subtyping of Y. enterocolitica. To this end, we developed a reference MS database library including 19 Y. enterocolitica (non-pathogenic biotype 1A and pathogenic biotypes 2 and 4) as well as 24 non-Y. enterocolitica strains, belonging to eleven different other Yersinia spp. The strains provided reproducible and unique mass spectra profiles covering a wide molecular mass range (2000 to 30,000 Da). Species-specific and biotype-specific biomarker protein mass patterns were determined for Y. enterocolitica. The defined biomarker mass patterns (SARAMIS SuperSpectrum™) were validated using 117 strains from various Y. enterocolitica bioserotypes in a blind-test. All strains were correctly identified and for all strains the mass spectrometry-based identification scheme yielded identical results compared to a characterization by a combination of biotyping and serotyping. Our study demonstrates that MALDI-TOF-MS is a reliable and powerful tool for the rapid identification of Y. enterocolitica strains to the species level and allows subtyping of strains to the biotype level. Copyright © 2011 Elsevier B.V. All rights reserved.

  16. Twin-Schnorr: A Security Upgrade for the Schnorr Identity-Based Identification Scheme

    Directory of Open Access Journals (Sweden)

    Ji-Jian Chin

    2015-01-01

    Full Text Available Most identity-based identification (IBI schemes proposed in recent literature are built using pairing operations. This decreases efficiency due to the high operation costs of pairings. Furthermore, most of these IBI schemes are proven to be secure against impersonation under active and concurrent attacks using interactive assumptions such as the one-more RSA inversion assumption or the one-more discrete logarithm assumption, translating to weaker security guarantees due to the interactive nature of these assumptions. The Schnorr-IBI scheme was first proposed through the Kurosawa-Heng transformation from the Schnorr signature. It remains one of the fastest yet most secure IBI schemes under impersonation against passive attacks due to its pairing-free design. However, when required to be secure against impersonators under active and concurrent attacks, it deteriorates greatly in terms of efficiency due to the protocol having to be repeated multiple times. In this paper, we upgrade the Schnorr-IBI scheme to be secure against impersonation under active and concurrent attacks using only the classical discrete logarithm assumption. This translates to a higher degree of security guarantee with only some minor increments in operational costs. Furthermore, because the scheme operates without pairings, it still retains its efficiency and superiority when compared to other pairing-based IBI schemes.

  17. Rapid Color Test Identification System for Screening of Counterfeit Fluoroquinolone

    Directory of Open Access Journals (Sweden)

    B. K. Singh

    2009-01-01

    Full Text Available The protocol of rapid identification system consists of three chemical color reactions; two group tests for fluoroquinolone class and a compound specific test each for norfloxacin, ciprofloxacin, gatifloxacin, ofloxacin, levofloxacin and sparfloxacin. The group color reactions are based on (a Oxidizing behavior of quinolone and (b Fluorine functional groups, both of which are characteristic of fluoroquinolone class. The compound specific color reactions are developed taking into consideration unique chemical behavior of each compound. The proposed chemical color tests have high selectivity⁄specificity, are ideal for screening purpose. The color of each test was defined by two standard color systems namely CIE lab and Munsell color. A suspected counterfeit tablet of any of the above mentioned drugs can be identified within 10-15 min using this rapid identification system.

  18. Provably secure identity-based identification and signature schemes from code assumptions

    Science.gov (United States)

    Zhao, Yiming

    2017-01-01

    Code-based cryptography is one of few alternatives supposed to be secure in a post-quantum world. Meanwhile, identity-based identification and signature (IBI/IBS) schemes are two of the most fundamental cryptographic primitives, so several code-based IBI/IBS schemes have been proposed. However, with increasingly profound researches on coding theory, the security reduction and efficiency of such schemes have been invalidated and challenged. In this paper, we construct provably secure IBI/IBS schemes from code assumptions against impersonation under active and concurrent attacks through a provably secure code-based signature technique proposed by Preetha, Vasant and Rangan (PVR signature), and a security enhancement Or-proof technique. We also present the parallel-PVR technique to decrease parameter values while maintaining the standard security level. Compared to other code-based IBI/IBS schemes, our schemes achieve not only preferable public parameter size, private key size, communication cost and signature length due to better parameter choices, but also provably secure. PMID:28809940

  19. Provably secure identity-based identification and signature schemes from code assumptions.

    Directory of Open Access Journals (Sweden)

    Bo Song

    Full Text Available Code-based cryptography is one of few alternatives supposed to be secure in a post-quantum world. Meanwhile, identity-based identification and signature (IBI/IBS schemes are two of the most fundamental cryptographic primitives, so several code-based IBI/IBS schemes have been proposed. However, with increasingly profound researches on coding theory, the security reduction and efficiency of such schemes have been invalidated and challenged. In this paper, we construct provably secure IBI/IBS schemes from code assumptions against impersonation under active and concurrent attacks through a provably secure code-based signature technique proposed by Preetha, Vasant and Rangan (PVR signature, and a security enhancement Or-proof technique. We also present the parallel-PVR technique to decrease parameter values while maintaining the standard security level. Compared to other code-based IBI/IBS schemes, our schemes achieve not only preferable public parameter size, private key size, communication cost and signature length due to better parameter choices, but also provably secure.

  20. Continuous-Flow Detector for Rapid Pathogen Identification

    Energy Technology Data Exchange (ETDEWEB)

    Barrett, Louise M. [Sandia National Lab. (SNL-CA), Livermore, CA (United States). Microfluidics; Skulan, Andrew J. [Sandia National Lab. (SNL-CA), Livermore, CA (United States). Microfluidics; Singh, Anup K. [Sandia National Lab. (SNL-CA), Livermore, CA (United States). Microfluidics; Cummings, Eric B. [Sandia National Lab. (SNL-CA), Livermore, CA (United States). Microfluidics; Fiechtner, Gregory J. [Sandia National Lab. (SNL-CA), Livermore, CA (United States). Microfluidics

    2006-09-01

    This report describes the continued development of a low-power, portable detector for the rapid identification of pathogens such as B. anthracis and smallpox. Based on our successful demonstration of the continuous filter/concentrator inlet, we believe strongly that the inlet section will enable differentiation between viable and non-viable populations, between types of cells, and between pathogens and background contamination. Selective, continuous focusing of particles in a microstream enables highly selective and sensitive identification using fluorescently labeled antibodies and other receptors such as peptides, aptamers, or small ligands to minimize false positives. Processes such as mixing and lysing will also benefit from the highly localized particle streams. The concentrator is based on faceted prisms to contract microfluidic flows while maintaining uniform flowfields. The resulting interfaces, capable of high throughput, serve as high-, low-, and band-pass filters to direct selected bioparticles to a rapid, affinity-based detection system. The proposed device is superior to existing array-based detectors as antibody-pathogen binding can be accomplished in seconds rather than tens of minutes or even hours. The system is being designed to interface with aerosol collectors under development by the National Laboratories or commercial systems. The focused stream is designed to be interrogated using diode lasers to differentiate pathogens by light scattering. Identification of particles is done using fluorescently labeled antibodies to tag the particles, followed by multiplexed laser-induced fluorescence (LIF) detection (achieved by labeling each antibody with a different dye).

  1. Method for Rapid Protein Identification in a Large Database

    Directory of Open Access Journals (Sweden)

    Wenli Zhang

    2013-01-01

    Full Text Available Protein identification is an integral part of proteomics research. The available tools to identify proteins in tandem mass spectrometry experiments are not optimized to face current challenges in terms of identification scale and speed owing to the exponential growth of the protein database and the accelerated generation of mass spectrometry data, as well as the demand for nonspecific digestion and post-modifications in complex-sample identification. As a result, a rapid method is required to mitigate such complexity and computation challenges. This paper thus aims to present an open method to prevent enzyme and modification specificity on a large database. This paper designed and developed a distributed program to facilitate application to computer resources. With this optimization, nearly linear speedup and real-time support are achieved on a large database with nonspecific digestion, thus enabling testing with two classical large protein databases in a 20-blade cluster. This work aids in the discovery of more significant biological results, such as modification sites, and enables the identification of more complex samples, such as metaproteomics samples.

  2. Rapid molecular identification of human taeniid cestodes by pyrosequencing approach.

    Directory of Open Access Journals (Sweden)

    Tongjit Thanchomnang

    Full Text Available Taenia saginata, T. solium, and T. asiatica are causative agents of taeniasis in humans. The difficulty of morphological identification of human taeniids can lead to misdiagnosis or confusion. To overcome this problem, several molecular methods have been developed, but use of these tends to be time-consuming. Here, a rapid and high-throughput pyrosequencing approach was developed for the identification of three human taeniids originating from various countries. Primers targeting the mitochondrial cytochrome c oxidase subunit 1 (cox1 gene of the three Taenia species were designed. Variations in a 26-nucleotide target region were used for identification. The reproducibility and accuracy of the pyrosequencing technology was confirmed by Sanger sequencing. This technique will be a valuable tool to distinguish between sympatric human taeniids that occur in Thailand, Asia and Pacific countries. This method could potentially be used for the molecular identification of the taeniid species that might be associated with suspicious cysts and lesions, or cyst residues in humans or livestock at the slaughterhouse.

  3. Rapid Molecular Identification of Human Taeniid Cestodes by Pyrosequencing Approach

    Science.gov (United States)

    Thanchomnang, Tongjit; Tantrawatpan, Chairat; Intapan, Pewpan M.; Sanpool, Oranuch; Janwan, Penchom; Lulitanond, Viraphong; Tourtip, Somjintana; Yamasaki, Hiroshi; Maleewong, Wanchai

    2014-01-01

    Taenia saginata, T. solium, and T. asiatica are causative agents of taeniasis in humans. The difficulty of morphological identification of human taeniids can lead to misdiagnosis or confusion. To overcome this problem, several molecular methods have been developed, but use of these tends to be time-consuming. Here, a rapid and high-throughput pyrosequencing approach was developed for the identification of three human taeniids originating from various countries. Primers targeting the mitochondrial cytochrome c oxidase subunit 1 (cox1) gene of the three Taenia species were designed. Variations in a 26-nucleotide target region were used for identification. The reproducibility and accuracy of the pyrosequencing technology was confirmed by Sanger sequencing. This technique will be a valuable tool to distinguish between sympatric human taeniids that occur in Thailand, Asia and Pacific countries. This method could potentially be used for the molecular identification of the taeniid species that might be associated with suspicious cysts and lesions, or cyst residues in humans or livestock at the slaughterhouse. PMID:24945530

  4. Rapid identification of cytokinins by an immunological method

    Energy Technology Data Exchange (ETDEWEB)

    Morris, R.O.; Jameson, P.E.; Morris, J.W. (Univ. of Missouri-Columbia (USA)); Laloue, M. (Centre Nationale de la Recherche Scientifique, Gif-sur-Yvette (France))

    1991-04-01

    A method for rapid identification of bacterial cytokinins has been developed in which cultures are fed ({sup 3}H)adenine, the cytokinins (including, {sup 3}H-labeled cytokinins) are isolated by immunoaffinity chromatography, and analyzed by HPLC with on-line scintillation counting. Analysis of Agrobacterium tumefaciens strains showed that some produced primarily trans-zeatin, whereas others produced primarily trans-zeatin riboside. Pseudomonas syringae pv savastanoi produced mixtures of transzeatin, dihydrozeatin, 1{double prime}-methyl-trans-zeatin riboside, and other unknown cytokinin-like substances. Corynebacterium fascians, produced cis-zeatin, isopentenyladenine and isopentenyladenosine. The technique is designed for qualitative rather than quantitative studies and allows ready identification of bacterial cytokinins. It may also have utility in the study of plant cytokinins if adequate incorporation of label into cytokinin precursor pools can be achieved.

  5. Use of a Generic Identification Scheme Connecting Events and Detector Description in the ATLAS Experiment

    CERN Document Server

    Arnault, C.

    2003-01-01

    High energy physics detectors can be described hierarchically from the different subsystems to their divisions in r, phi, theta and to the individual readout channels. An identification schema that follows the logical decomposition of the ATLAS detector has been introduced allowing identification of individual readout channels as well as other parts of the detector, in particular detector elements. These identifiers provide a sort of ?glue? allowing, for example, the connection of raw event data to their detector description for position calculation or alignment corrections, as well as fast access to subsets of the event data for event trigger selection. There are two important requirements on the software to support such an identification scheme. First is the possibility to formally specify these identifiers in terms of their structure and allowed values. And second is to generate different forms of the identifiers optimised in terms of access efficiency to information content, compactness or search key effi...

  6. Multi-stage identification scheme for detecting damage in structures under ambient excitations

    Science.gov (United States)

    Bao, Chunxiao; Hao, Hong; Li, Zhong-Xian

    2013-04-01

    Structural damage identification methods are critical to the successful application of structural health monitoring (SHM) systems to civil engineering structures. The dynamic response of civil engineering structures is usually characterized by high nonlinearity and non-stationarity. Accordingly, an improved Hilbert-Huang transform (HHT) method which is adaptive, output-only and applicable to system identification of in-service structures under ambient excitations is developed in this study. Based on this method, a multi-stage damage detection scheme including the detection of damage occurrence, damage existence, damage location and the estimation of damage severity is developed. In this scheme, the improved HHT method is used to analyse the structural acceleration response, the obtained instantaneous frequency detects the instant of damage occurrence, the instantaneous phase is sensitive to minor damage and provides reliable damage indication, and the damage indicator developed based on statistical analysis of the Hilbert marginal spectrum detects damage locations. Finally, the response sampled at the detected damage location is continuously analysed to estimate the damage severity. Numerical and experimental studies of frame structures under ambient excitations are performed. The results demonstrate that this scheme accomplishes the above damage detection functions within one flow. It is robust, time efficient, simply implemented and applicable to the real-time SHM of in-service structures.

  7. Rapid identification of Sporothrix schenckii in biopsy tissue by PCR.

    Science.gov (United States)

    Liu, X; Zhang, Z; Hou, B; Wang, D; Sun, T; Li, F; Wang, H; Han, S

    2013-12-01

    The dimorphic fungus Sporothrix schenckii is the etiological agent of sporotrichosis, an important cutaneous mycosis with a worldwide distribution. At present, it is challenging to rapidly discover and identify Sporothrix schenckii in biopsy tissues nowadays. To explore new methods for rapid diagnosis of sporotrichosis. We screened specific primers for Sporothrix schenckii using 50 clinical isolates from patients with sporotrichosis. DNA was extracted from the lesions of 30 cases of clinically suspected sporotrichosis using the Graham s method of CTAB and amplified by PCR using the screened specific primers. The primer S2-R2 was applicable for the identification of S. schenckii from different geographic areas and clinical types with high specificity and sensitivity. Twenty-five out of the thirty cases (83.3%) amplified using the primer S2-R2 showed positive bands. Further positive bands were observed in 95.6% of cases tested positive by fungal culture. Using the PCR technique and specific primers, we developed a new diagnostic method that can rapidly diagnose sporotrichosis with tissues obtained from clinical biopsies. © 2012 The Authors. Journal of the European Academy of Dermatology and Venereology © 2012 European Academy of Dermatology and Venereology.

  8. Elucidation of molecular kinetic schemes from macroscopic traces using system identification.

    Directory of Open Access Journals (Sweden)

    Miguel Fribourg

    2017-02-01

    Full Text Available Overall cellular responses to biologically-relevant stimuli are mediated by networks of simpler lower-level processes. Although information about some of these processes can now be obtained by visualizing and recording events at the molecular level, this is still possible only in especially favorable cases. Therefore the development of methods to extract the dynamics and relationships between the different lower-level (microscopic processes from the overall (macroscopic response remains a crucial challenge in the understanding of many aspects of physiology. Here we have devised a hybrid computational-analytical method to accomplish this task, the SYStems-based MOLecular kinetic scheme Extractor (SYSMOLE. SYSMOLE utilizes system-identification input-output analysis to obtain a transfer function between the stimulus and the overall cellular response in the Laplace-transformed domain. It then derives a Markov-chain state molecular kinetic scheme uniquely associated with the transfer function by means of a classification procedure and an analytical step that imposes general biological constraints. We first tested SYSMOLE with synthetic data and evaluated its performance in terms of its rate of convergence to the correct molecular kinetic scheme and its robustness to noise. We then examined its performance on real experimental traces by analyzing macroscopic calcium-current traces elicited by membrane depolarization. SYSMOLE derived the correct, previously known molecular kinetic scheme describing the activation and inactivation of the underlying calcium channels and correctly identified the accepted mechanism of action of nifedipine, a calcium-channel blocker clinically used in patients with cardiovascular disease. Finally, we applied SYSMOLE to study the pharmacology of a new class of glutamate antipsychotic drugs and their crosstalk mechanism through a heteromeric complex of G protein-coupled receptors. Our results indicate that our methodology

  9. Emerging technologies for rapid identification of bloodstream pathogens.

    Science.gov (United States)

    Kothari, Atul; Morgan, Margie; Haake, David A

    2014-07-15

    Technologies for rapid microbial identification are poised to revolutionize clinical microbiology and enable informed decision making for patients with life-threatening bloodstream infections. Species identification of microorganisms in positive blood cultures can be performed in minutes using commercial fluorescence in situ hybridization tests or mass spectroscopy. Microorganisms in positive blood cultures can also be identified within 1-2.5 hours using automated polymerase chain reaction-based systems that can also detect selected antibiotic resistance markers, such as methicillin resistance. When combined with antibiotic stewardship programs, these approaches improve clinical outcomes and reduce healthcare expenditures. Tests for direct detection in whole blood samples are highly desirable because of their potential to identify bloodstream pathogens without waiting 1-2 days for blood cultures to become positive. However, results for pathogen detection in whole blood do not overlap with those of conventional blood culture techniques and we are still learning how best to use these approaches. © The Author 2014. Published by Oxford University Press on behalf of the Infectious Diseases Society of America. All rights reserved. For Permissions, please e-mail: journals.permissions@oup.com.

  10. Rapid identification of non-sporing anaerobes using nuclear magnetic resonance spectroscopy and an identification strategy

    Directory of Open Access Journals (Sweden)

    Menon S

    2007-01-01

    Full Text Available Purpose: The non-sporing anaerobes cause a wide spectrum of infections. They are difficult to culture and their identification is tedious and time-consuming. Rapid identification of anaerobes is highly desirable. Towards this end, the potential of nuclear magnetic resonance (NMR spectroscopy for providing a fingerprint within the proton spectrum of six genera belonging to anaerobes reflecting their characteristic metabolites has been investigated. Methods: NMR analysis was carried out using Mercury plus Varian 300 MHz (7.05 T NMR spectrophotometer on six different anaerobes. These included Bacteroides fragilis, Prevotella melaninogenica, Prevotella denticola, Fusobacterium necrophorum, Peptococcus niger and Peptostreptococcus spp. After the NMR analysis (256/512 scans, the different peaks were noted. The eight pus specimens, which yielded pure culture of anaerobe, also were analysed similarly. Results: The major resonances of multiplex of amino acids/lipid at 0.9 ppm along with lactate/lipid at 1.3 ppm, acetate at 1.92 ppm and multiplex of lysine at 3.0 ppm remained constant to label the organism as an anaerobe. There was a difference found in the MR spectra of different genera and species. A simple algorithm was developed for the identification of the six different anaerobes studied. The MR spectra of the pure culture of the organism matched the MR spectra of pus from which the organism was isolated. Conclusions: MR-based identification was of value in the identification of anaerobes. However, a larger database of the peaks produced by anaerobes needs to be created for identification of all genera and species. It could then have the potential of diagnosing an anaerobic infection in vivo and thus expedite management of deep-seated abscesses.

  11. [Rapid identification of microorganisms using MALDI-TOF mass spectrometry].

    Science.gov (United States)

    Sogawa, Kazuyuki; Watanabe, Masaharu; Nomura, Fumio

    2013-01-01

    In a clinical diagnostic microbiology laboratory, the current method of identifying bacterial isolates is based mainly on phenotypic characteristics, such as the growth pattern on different media, colony morphology, Gram stain, and various biochemical reactions. These techniques collectively allow high-level accuracy in identifying most bacterial isolates, but they are costly and time-consuming. In our clinical microbiology laboratory, we prospectively assessed the ability of matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS) to identify bacterial strains that were routinely isolated from clinical samples. Bacterial colonies obtained from a total of 468 strains of 92 bacterial species isolated at the Department of Clinical Laboratory at Chiba University were directly placed on target MALDI plates, followed by the addition of CHCA matrix solution. The plates were then subjected to MALDI-TOF MS measurement, and the microorganisms were identified by pattern matching by the libraries in the BioTyper 2.0 software. The identification rates at species and genus levels were 91.7% (429/468) and 97.0% (454/468), respectively. MALDI-TOF MS is a rapid, simple, and high-throughput proteomic technique for the identification of a variety of bacterial species. Since colony to colony differences and the effects of culture duration on the results are minimal, it can be implemented in a conventional laboratory setting. Although for some pathogens, the preanalytic processes should be refined and current database should be improved to obtain more accurate results, the MALDI-TOF MS-based method generally performs as well as the conventional methods and is a promising technology in clinical laboratories.

  12. Artificial neural network based fault identification scheme implementation for a three-phase induction motor.

    Science.gov (United States)

    Kolla, Sri R; Altman, Shawn D

    2007-04-01

    This paper presents results from the implementation and testing of a PC based monitoring and fault identification scheme for a three-phase induction motor using artificial neural networks (ANNs). To accomplish the task, a hardware system is designed and built to acquire three-phase voltages and currents from a 1/3 HP squirrel-cage, three-phase induction motor. A software program is written to read the voltages and currents, which are first used to train a feed-forward neural network structure using the JavaNNS program. The trained network is placed in a LabVIEW based program formula node that monitors the voltages and currents online and displays the fault conditions and turns the motor off. The complete system is successfully tested in real time by creating different faults on the motor.

  13. Rapid and generic identification of influenza A and other respiratory viruses with mass spectrometry

    NARCIS (Netherlands)

    Majchrzykiewicz-Koehorst, J.A.; Heikens, E.; Trip, H.; Hulst, A.G.; Jong, A.L. de; Viveen, M.C.; Sedee, N.J.A.; van der Plas, J.; Coenjaerts, F.E.J.; Paauw, A.

    2015-01-01

    The rapid identification of existing and emerging respiratory viruses is crucial in combating outbreaks and epidemics. Matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) is a rapid and reliable identification method in bacterial diagnostics, but has not been

  14. Rapid identification of pathogens in blood cultures with a modified fluorescence in situ hybridization assay

    NARCIS (Netherlands)

    Peters, Remco P. H.; van Agtmael, Michiel A.; Simoons-Smit, Alberdina M.; Danner, Sven A.; Vandenbroucke-Grauls, Christina M. J. E.; Savelkoul, Paul H. M.

    2006-01-01

    We evaluated a modified fluorescence in situ hybridization (FISH) assay for rapid ( <1 h) identification of microorganisms in growth-positive blood cultures. The results were compared to those of the standard FISH technique and conventional culturing. The rapid identification of microorganisms with

  15. Rapid analysis for the identification of the seagrass Halophila ovalis ...

    African Journals Online (AJOL)

    Maslin

    2015-02-25

    Feb 25, 2015 ... conventional identification keys alone: i) the variability of morphological characteristics and ii) lack of needed morphological ... Key words: DNA barcoding, Halophila ovalis, rbcL, trnH-psbA, species identification. INTRODUCTION ..... of lignified tissue, the structure of seagrasses is flexible and vulnerable to.

  16. Rapid method for identification of transgenic fish zygosity

    Directory of Open Access Journals (Sweden)

    . Alimuddin

    2007-07-01

    Full Text Available Identification of zygosity in transgenik fish is normally achieved by PCR analysis with genomic DNA template extracted from the tissue of progenies which are derived by mating the transgenic fish and wild-type counterpart.  This method needs relatively large amounts of fish material and is time- and labor-intensive. New approaches addressing this problem could be of great help for fish biotechnologists.  In this experiment, we applied a quantitative real-time PCR (qr-PCR method to analyze zygosity in a stable line of transgenic zebrafish (Danio rerio carrying masu salmon, Oncorhynchus masou D6-desaturase-like gene. The qr-PCR was performed using iQ SYBR Green Supermix in the iCycler iQ Real-time PCR Detection System (Bio-Rad Laboratories, USA.  Data were analyzed using the comparative cycle threshold method.  The results demonstrated a clear-cut identification of all transgenic fish (n=20 classified as a homozygous or heterozygous.  Mating of those fish with wild-type had revealed transgene transmission to the offspring following expected Mendelian laws. Thus, we found that the qTR-PCR to be effective for a rapid and precise determination of zygosity in transgenic fish. This technique could be useful in the establishment of breeding programs for mass transgenic fish production and in experiments in which zygosity effect could have a functional impact. Keywords: quantitative real-time PCR; zygosity; transgenic fish; mass production   ABSTRAK Identifikasi sigositas ikan transgenik biasanya dilakukan menggunakan analisa PCR dengan cetakan DNA genomik yang diekstraksi dari jaringan ikan hasil persilangan antara ikan transgenik dan ikan normal.   Metode ini memerlukan ikan dalam jumlah yang banyak, dan juga waktu serta tenaga.  Pendekatan baru untuk mengatasi masalah tersebut akan memberikan manfaat besar kepada peneliti bioteknologi perikanan.  Pada penelitian ini, kami menggunakan metode PCR real-time kuantitatif (krt-PCR untuk

  17. Parameter Identification and Control Scheme for Monitoring Automatic Thickness Control System with Measurement Delay

    Directory of Open Access Journals (Sweden)

    Xu Yang

    2017-01-01

    Full Text Available The thickness of the steel strip is an important indicator of the overall strip quality. Deviations in thickness are primarily controlled using the automatic gauge control (AGC system of each rolling stand. At the last stand, the monitoring AGC system is usually used, where the deviations in thickness can be directly measured by the X-ray thickness gauge device and used as the input to the AGC system. However, due to the physical distance between the thickness detection device and the rolling stand, time delay is unavoidably present in the thickness control loop, which can affect control performance and lead to system oscillations. Furthermore, the parameters of the system can change due to perturbations from external disturbances. Therefore, this paper proposes an identification and control scheme for monitoring AGC system that can handle time delay and parameter uncertainty. The cross-correlation function is used to estimate the time delay of the system, while the system parameters are identified using a recursive least squares method. The time delay and parameter estimates are then further refined using the Levenberg-Marquardt algorithm, so as to provide the most accurate parameter estimates for the complete system. Simulation results show that, compared with the standard Proportion Integration Differentiation (PID controller approach, the proposed approach is not affected by changes in the time delay and parameter uncertainties.

  18. Rapid identification of single microbes by various Raman spectroscopic techniques

    Science.gov (United States)

    Rösch, Petra; Harz, Michaela; Schmitt, Michael; Peschke, Klaus-Dieter; Ronneberger, Olaf; Burkhardt, Hans; Motzkus, Hans-Walter; Lankers, Markus; Hofer, Stefan; Thiele, Hans; Popp, Jürgen

    2006-02-01

    A fast and unambiguous identification of microorganisms is necessary not only for medical purposes but also in technical processes such as the production of pharmaceuticals. Conventional microbiological identification methods are based on the morphology and the ability of microbes to grow under different conditions on various cultivation media depending on their biochemical properties. These methods require pure cultures which need cultivation of at least 6 h but normally much longer. Recently also additional methods to identify bacteria are established e.g. mass spectroscopy, polymerase chain reaction (PCR), flow cytometry or fluorescence spectroscopy. Alternative approaches for the identification of microorganisms are vibrational spectroscopic techniques. With Raman spectroscopy a spectroscopic fingerprint of the microorganisms can be achieved. Using UV-resonance Raman spectroscopy (UVRR) macromolecules like DNA/RNA and proteins are resonantly enhanced. With an excitation wavelength of e.g. 244 nm it is possible to determine the ratio of guanine/cytosine to all DNA bases which allows a genotypic identification of microorganisms. The application of UVRR requires a large amount of microorganisms (> 10 6 cells) e.g. at least a micro colony. For the analysis of single cells micro-Raman spectroscopy with an excitation wavelength of 532 nm can be used. Here, the obtained information is from all type of molecules inside the cells which lead to a chemotaxonomic identification. In this contribution we show how wavelength dependent Raman spectroscopy yields significant molecular information applicable for the identification of microorganisms on a single cell level.

  19. Rapid Electrochemical Detection and Identification of Microbiological and Chemical Contaminants for Manned Spaceflight Project

    Data.gov (United States)

    National Aeronautics and Space Administration — A great deal of effort has gone into the development of point-of-use methods to meet the challenge of rapid bacterial identification for both environmental...

  20. Rapid identification of nine species of diphyllobothriidean tapeworms by pyrosequencing

    Science.gov (United States)

    Thanchomnang, Tongjit; Tantrawatpan, Chairat; Intapan, Pewpan M.; Sanpool, Oranuch; Lulitanond, Viraphong; Tourtip, Somjintana; Yamasaki, Hiroshi; Maleewong, Wanchai

    2016-01-01

    The identification of diphyllobothriidean tapeworms (Cestoda: Diphyllobothriidea) that infect humans and intermediate/paratenic hosts is extremely difficult due to their morphological similarities, particularly in the case of Diphyllobothrium and Spirometra species. A pyrosequencing method for the molecular identification of pathogenic agents has recently been developed, but as of yet there have been no reports of pyrosequencing approaches that are able to discriminate among diphyllobothriidean species. This study, therefore, set out to establish a pyrosequencing method for differentiating among nine diphyllobothriidean species, Diphyllobothrium dendriticum, Diphyllobothrium ditremum, Diphyllobothrium latum, Diphyllobothrium nihonkaiense, Diphyllobothrium stemmacephalum, Diplogonoporus balaenopterae, Adenocephalus pacificus, Spirometra decipiens and Sparganum proliferum, based on the mitochondrial cytochrome c oxidase subunit 1 (cox1) gene as a molecular marker. A region of 41 nucleotides in the cox1 gene served as a target, and variations in this region were used for identification using PCR plus pyrosequencing. This region contains nucleotide variations at 12 positions, which is enough for the identification of the selected nine species of diphyllobothriidean tapeworms. This method was found to be a reliable tool not only for species identification of diphyllobothriids, but also for epidemiological studies of cestodiasis caused by diphyllobothriidean tapeworms at public health units in endemic areas. PMID:27853295

  1. Problems with rapid agglutination methods for identification of Staphylococcus aureus when Staphylococcus saprophyticus is being tested.

    OpenAIRE

    Gregson, D B; Low, D E; Skulnick, M; Simor, A E

    1988-01-01

    Six rapid agglutination tests for identification of Staphylococcus aureus were evaluated by using 62 strains of S. aureus, 63 strains of S. saprophyticus, and 67 strains of other coagulase-negative staphylococci. S. saprophyticus was responsible for 19 of 26 false-positive results and 20 uninterpretable reactions. Thus, urinary staphylococcal isolates that are positive by rapid agglutination tests may require other confirmatory tests for the identification of possible S. saprophyticus.

  2. Rapid identification of pathogens from pediatric blood cultures by use of the FilmArray blood culture identification panel.

    Science.gov (United States)

    Zheng, Xiaotian; Polanco, Wanda; Carter, Donna; Shulman, Stanford

    2014-12-01

    The performance of the FilmArray blood culture identification (BCID) panel has been studied in adult patients. We describe here an evaluation of this assay for the rapid identification of pathogens in Bactec Peds Plus/F and Bactec standard anaerobic/F bottles that contained blood samples from pediatric patients at a tertiary care children's hospital. Copyright © 2014, American Society for Microbiology. All Rights Reserved.

  3. Rapid and Accurate Identification of Adulterants via an Electronic Nose and DNA Identification Platform: Identification of Fake Velvet Antlers as an Example

    OpenAIRE

    Guojie Xu; Chunsheng Liu; Caili Liao; Xiaolei Ren; Xinyue Zhang; Xiaorui Fu; Xueyong Wang

    2016-01-01

    Background. Adulterants in Chinese medicines had always affected the efficacy of Chinese medicines and resulted in safety problems in drug use. We aimed to take the identification of fake velvet antlers as an example for establishment of a rapid and accurate identification platform for modern pharmaceutical companies and markets of Chinese medicine. Methods. In this study, we developed a novel electronic nose and DNA identification platform for identifying fake velvet antlers. Electronic nose...

  4. Rapid identification of the medicinal plant Taraxacum formosanum ...

    African Journals Online (AJOL)

    Administrator

    2011-06-06

    Jun 6, 2011 ... College J. 8: 35-46. Xue CY, Li DZ, Lu JM, Yang JB, Liu JQ (2006). Molecular authentication of the traditional Tibetan medical plant Swertia mussotii. Planta Med. 72: 721-726. Yuan CC (2001). Textual research of material medica Taraxacum mongolicum and varietal identification. Chinese Wild Plant Res.

  5. Rapid identification of the medicinal plant Taraxacum formosanum ...

    African Journals Online (AJOL)

    Administrator

    2011-06-06

    Jun 6, 2011 ... Original identification of medicinal plants is essential for quality control. In this study, the internal transcribed spacer 2 (ITS2) nuclear ribosomal DNA served as a DNA barcode and was amplified by allele-specific PCR. This approach was exploited to differentiate Taraxacum formosanum from five.

  6. Non-intrusive schemes for speed and axle identification in bridge-weigh-in-motion systems

    Science.gov (United States)

    Kalhori, Hamed; Makki Alamdari, Mehrisadat; Zhu, Xinqun; Samali, Bijan; Mustapha, Samir

    2017-02-01

    Bridge weigh-in-motion (BWIM) is an approach through which the axle and gross weight of trucks travelling at normal highway speed are identified using the response of an instrumented bridge. The vehicle speed, the number of axles, and the axle spacing are crucial parameters, and are required to be determined in the majority of BWIM algorithms. Nothing-on-the-road (NOR) strategy suggests using the strain signals measured at some particular positions underneath the deck or girders of a bridge to obtain this information. The objective of this research is to present a concise overview of the challenges of the current non-intrusive schemes for speed and axle determination through bending-strain and shear-strain based approaches. The problem associated with the global bending-strain responses measured at quarter points of span is discussed and a new sensor arrangement is proposed as an alternative. As for measurement of local responses rather than the global responses, the advantage of shear strains over bending strains is presented. However, it is illustrated that shear strains at quarter points of span can only provide accurate speed estimation but fail to detect the correct number of axles. As a remedy, it is demonstrated that, even for closely-spaced axles, the shear strain at the beginning of the bridge is capable of reliably identifying the number of axles. In order to provide a fully automated speed and axle identification system, appropriate signal processing including low-pass filtering and wavelet transforms are applied to the raw time signals. As case studies, the results of experimental testing in laboratory and on a real bridge are presented.

  7. Rapid identification of bio-molecules applied for detection of biosecurity agents using rolling circle amplification.

    Directory of Open Access Journals (Sweden)

    Jenny Göransson

    Full Text Available Detection and identification of pathogens in environmental samples for biosecurity applications are challenging due to the strict requirements on specificity, sensitivity and time. We have developed a concept for quick, specific and sensitive pathogen identification in environmental samples. Target identification is realized by padlock- and proximity probing, and reacted probes are amplified by RCA (rolling-circle amplification. The individual RCA products are labeled by fluorescence and enumerated by an instrument, developed for sensitive and rapid digital analysis. The concept is demonstrated by identification of simili biowarfare agents for bacteria (Escherichia coli and Pantoea agglomerans and spores (Bacillus atrophaeus released in field.

  8. A Novel, Simplified Scheme for Plastics Identification: "JCE" Classroom Activity 104

    Science.gov (United States)

    Harris, Mary E.; Walker, Barbara

    2010-01-01

    In this Activity, students identify samples of seven types of recyclable plastic by using a flowchart scheme. The flowchart procedure includes making density comparisons of the plastic samples in water and alcohol and observing physical changes of plastic samples subjected to boiling water temperatures and exposure to acetone. This scheme is…

  9. An on-line equivalent system identification scheme for adaptive control. Ph.D. Thesis - Stanford Univ.

    Science.gov (United States)

    Sliwa, S. M.

    1984-01-01

    A prime obstacle to the widespread use of adaptive control is the degradation of performance and possible instability resulting from the presence of unmodeled dynamics. The approach taken is to explicitly include the unstructured model uncertainty in the output error identification algorithm. The order of the compensator is successively increased by including identified modes. During this model building stage, heuristic rules are used to test for convergence prior to designing compensators. Additionally, the recursive identification algorithm as extended to multi-input, multi-output systems. Enhancements were also made to reduce the computational burden of an algorithm for obtaining minimal state space realizations from the inexact, multivariate transfer functions which result from the identification process. A number of potential adaptive control applications for this approach are illustrated using computer simulations. Results indicated that when speed of adaptation and plant stability are not critical, the proposed schemes converge to enhance system performance.

  10. A rapid, one step molecular identification of Trichoderma citrinoviride and Trichoderma reesei.

    Science.gov (United States)

    Saroj, Dina B; Dengeti, Shrinivas N; Aher, Supriya; Gupta, Anil K

    2015-06-01

    Trichoderma species are widely used as production hosts for industrial enzymes. Identification of Trichoderma species requires a complex molecular biology based identification involving amplification and sequencing of multiple genes. Industrial laboratories are required to run identification tests repeatedly in cell banking procedures and also to prove absence of production host in the product. Such demands can be fulfilled by a brief method which enables confirmation of strain identity. This communication describes one step identification method for two common Trichoderma species; T. citrinoviride and T. reesei, based on identification of polymorphic region in the nucleotide sequence of translation elongation factor 1 alpha. A unique forward primer and common reverse primer resulted in 153 and 139 bp amplicon for T. citrinoviride and T. reesei, respectively. Simplification was further introduced by using mycelium as template for PCR amplification. Method described in this communication allows rapid, one step identification of two Trichoderma species.

  11. Rapid identification of Mycobacterium species by lectin agglutination.

    Science.gov (United States)

    Athamna, Abed; Cohen, Dani; Athamna, Muhammad; Ofek, Itzhak; Stavri, Henriette

    2006-05-01

    The purpose of the present study is to explore the possibility that plant lectins can be used for the development of rapid and inexpensive technique for differentiation of mycobacterial species. The method is based on interaction between mycobacteria and lectins as visualized by agglutination in a microtiter plate. We employed 18 mycobacterium species and determined the minimal lectin concentration (MLC) of 23 different lectins. For some of the bacteria as a high as 1000 microg/ml of one or more lectins were required to induce agglutination, while for other strains as low as 1.95 microg/ml of the lectin were needed. A unique pattern of agglutination was observed for each species over a range of 62-1000 microg/ml lectin concentrations. There were little or no variations in MLC within strains (intraspecies) of each of two species tested. In contrast, there were marked interspecies variations in MLC. Analysis of the MLC showed that the highest score of interspecies differences with 23 lectins was obtained at 125 microg/ml lectin concentration. At this concentration it was found that the pattern of agglutinations with only two lectins was sufficient to differentiate mycobacterium species from each other. Because the bacteria-lectin interaction is adaptable to various methods of visualization, our findings may set the stage for developing a rapid and reliable tool to differentiate mycobacterium species.

  12. Small acid soluble proteins for rapid spore identification.

    Energy Technology Data Exchange (ETDEWEB)

    Branda, Steven S.; Lane, Todd W.; VanderNoot, Victoria A.; Jokerst, Amanda S.

    2006-12-01

    This one year LDRD addressed the problem of rapid characterization of bacterial spores such as those from the genus Bacillus, the group that contains pathogenic spores such as B. anthracis. In this effort we addressed the feasibility of using a proteomics based approach to spore characterization using a subset of conserved spore proteins known as the small acid soluble proteins or SASPs. We proposed developing techniques that built on our previous expertise in microseparations to rapidly characterize or identify spores. An alternative SASP extraction method was developed that was amenable to both the subsequent fluorescent labeling required for laser-induced fluorescence detection and the low ionic strength requirements for isoelectric focusing. For the microseparations, both capillary isoelectric focusing and chip gel electrophoresis were employed. A variety of methods were evaluated to improve the molecular weight resolution for the SASPs, which are in a molecular weight range that is not well resolved by the current methods. Isoelectric focusing was optimized and employed to resolve the SASPs using UV absorbance detection. Proteomic signatures of native wild type Bacillus spores and clones genetically engineered to produce altered SASP patterns were assessed by slab gel electrophoresis, capillary isoelectric focusing with absorbance detection as well as microchip based gel electrophoresis employing sensitive laser-induced fluorescence detection.

  13. [Rapid identification of Coix seed varieties by near infrared spectroscopy].

    Science.gov (United States)

    Liu, Xing; Mao, Dan-Zhuo; Wang, Zheng-Wu; Yang, Yong-Jian

    2014-05-01

    Unsupervised learning algorithm-principal component analysis (PCA), and supervised learning algorithm-learning vector quantization (LVQ) neural network and support vector machine (SVM) were used to carry out qualitative discriminant analysis of different varieties of coix seed from different regions. Since nutrient compositions of different varieties coix seed samples from different origins were complex and the contents were similar, characteristic variables of two kinds of coix seed were alike, the scores plot of their principal components seriously overlapped and the categories of coix seed were difficult to distinguish While satisfactory results were obtained by LVQ neural network and SVM. The accuracy of LVQ neural network prediction is 90. 91%, while the classification accuracy of SVM, whose penalty parameter and kernel function parameter were optimized, can be up to 100%. The results show that NIRS combined with chemometrics can be used as a rapid, nondestructive and reliable method to identify coix seed varieties and provide technical reference for market regulation.

  14. Rapid identification of mycobacteria and rapid detection of drug resistance in Mycobacterium tuberculosis in cultured isolates and in respiratory specimens.

    Science.gov (United States)

    Yam, Wing-Cheong; Siu, Kit-Hang Gilman

    2013-01-01

    Recent advances in molecular biology and better understanding of the genetic basis of drug resistance have allowed rapid identification of mycobacteria and rapid detection of drug resistance of Mycobacterium tuberculosis present in cultured isolates or in respiratory specimens. In this chapter, several simple nucleic acid amplification-based techniques are introduced as molecular approach for clinical diagnosis of tuberculosis. A one-tube nested IS6110-based polymerase chain reaction (PCR) is used for M. tuberculosis complex identification; the use of a multiplex allele-specific PCR is demonstrated to detect the isoniazid resistance; PCR-sequencing assays are applied for rifampicin and ofloxacin resistance detection and 16S rDNA sequencing is utilized for identification of mycobacterial species from cultures of acid fast bacilli (AFB). Despite the high specificity and sensitivity of the molecular techniques, mycobacterial culture remains the "Gold Standard" for tuberculosis diagnosis. Negative results of molecular tests never preclude the infection or the presence of drug resistance. These technological advancements are, therefore, not intended to replace the conventional tests, but rather have major complementary roles in tuberculosis diagnosis.

  15. Rapidly Learned Identification of Epileptic Seizures from Sonified EEG

    Directory of Open Access Journals (Sweden)

    Psyche eLoui

    2014-10-01

    Full Text Available Sonification refers to a process by which data are converted into sound, providing an auditory alternative to visual display. Currently, the prevalent method for diagnosing seizures in epilepsy is by visually reading a patient’s electroencephalogram (EEG. However, sonification of the EEG data provides certain advantages due to the nature of human auditory perception. We hypothesized that human listeners will be able to identify seizures from EEGs using the auditory modality alone, and that accuracy of seizure identification will increase after a short training session. Here we describe an algorithm we have used to sonify EEGs of both seizure and non-seizure activity, followed by a training study in which subjects listened to short clips of sonified EEGs and determine whether each clip was of seizure or normal activity, both before and after a short training session. Results show that before training subjects performed at chance level in differentiating seizures vs. non-seizures, but there was a significant improvement of accuracy after the training session. After training, subjects successfully distinguished seizures from non-seizures using the auditory modality alone. Further analyses using signal detection theory demonstrated improvement in sensitivity and reduction in response bias as a result of training. This study demonstrates the potential of sonified EEGs to be used for the detection of seizures. Future studies will attempt to increase accuracy using novel training and sonification modifications, with the goals of managing, predicting, and ultimately controlling seizures using sonification as a possible biofeedback-based intervention for epilepsy.

  16. Rapidly learned identification of epileptic seizures from sonified EEG.

    Science.gov (United States)

    Loui, Psyche; Koplin-Green, Matan; Frick, Mark; Massone, Michael

    2014-01-01

    Sonification refers to a process by which data are converted into sound, providing an auditory alternative to visual display. Currently, the prevalent method for diagnosing seizures in epilepsy is by visually reading a patient's electroencephalogram (EEG). However, sonification of the EEG data provides certain advantages due to the nature of human auditory perception. We hypothesized that human listeners will be able to identify seizures from EEGs using the auditory modality alone, and that accuracy of seizure identification will increase after a short training session. Here, we describe an algorithm that we have used to sonify EEGs of both seizure and non-seizure activity, followed by a training study in which subjects listened to short clips of sonified EEGs and determined whether each clip was of seizure or normal activity, both before and after a short training session. Results show that before training subjects performed at chance level in differentiating seizures from non-seizures, but there was a significant improvement of accuracy after the training session. After training, subjects successfully distinguished seizures from non-seizures using the auditory modality alone. Further analyses using signal detection theory demonstrated improvement in sensitivity and reduction in response bias as a result of training. This study demonstrates the potential of sonified EEGs to be used for the detection of seizures. Future studies will attempt to increase accuracy using novel training and sonification modifications, with the goals of managing, predicting, and ultimately controlling seizures using sonification as a possible biofeedback-based intervention for epilepsy.

  17. Rapid Identification of Aldose Reductase Inhibitory Compounds from Perilla frutescens

    Directory of Open Access Journals (Sweden)

    Ji Hun Paek

    2013-01-01

    Full Text Available The ethyl acetate (EtOAc soluble fraction of methanol extracts of Perilla frutescens (P. frutescens inhibits aldose reductase (AR, the key enzyme in the polyol pathway. Our investigation of inhibitory compounds from the EtOAc soluble fraction of P. frutescens was followed by identification of the inhibitory compounds by a combination of HPLC microfractionation and a 96-well enzyme assay. This allowed the biological activities to be efficiently matched with selected HPLC peaks. Structural analyses of the active compounds were performed by LC-MSn. The main AR inhibiting compounds were tentatively identified as chlorogenic acid and rosmarinic acid by LC-MSn. A two-step high speed counter current chromatography (HSCCC isolation method was developed with a solvent system of n-hexane-ethyl acetate-methanol-water at 1.5 : 5 : 1 : 5, v/v and 3 : 7 : 5 : 5, v/v. The chemical structures of the isolated compounds were determined by 1H- and 13C-nuclear magnetic resonance spectrometry (NMR. The main compounds inhibiting AR in the EtOAc fraction of methanol extracts of P. frutescens were identified as chlorogenic acid (2 (IC50 = 3.16 μM, rosmarinic acid (4 (IC50 = 2.77 μM, luteolin (5 (IC50 = 6.34 μM, and methyl rosmarinic acid (6 (IC50 = 4.03 μM.

  18. Rapid identification of antibiotic resistance using droplet microfluidics.

    Science.gov (United States)

    Keays, Marie C; O'Brien, Mark; Hussain, Anam; Kiely, Patrick A; Dalton, Tara

    2016-04-02

    Culturing bacteria and monitoring bacterial cell growth is a critical issue when dealing with patients who present with bacterial infections. One of the main challenges that arises is the time taken to identify the particular strain of bacteria and consequently, decide the correct treatment. In the majority of cases, broad spectrum antibiotics are used to target infections when a narrow spectrum drug would be more appropriate. The efficient monitoring of bacterial growth and potential antibiotic resistance is necessary to identify the best treatment options for patients. Minturising the reactions into microfluidic droplets offers a novel method to rapidy analyze bacteria. Microfluidics facilitates low volume reactions that provide a unique system where each droplet reaction acts as an individual bioreactor. Here, we designed and built a novel platform that allowed us to create and monitor E.coli microfluidic droplet cultures. Optical capacity was built in and measurements of bacterial cultures were captured facilitating the continuous monitoring of individual reactions. The capacity of the instrument was demonstrated by the application of treatments to both bacteria and drug resistant strains of bacteria. We were able to detect responses within one hour in the droplet cultures, demonstrating the capacity of this workflow to the culture and rapid characterization of bacterial strains.

  19. Mycobacteria mobility shift assay: a method for the rapid identification of Mycobacterium tuberculosis and nontuberculous mycobacteria

    Directory of Open Access Journals (Sweden)

    Letícia Muraro Wildner

    2014-06-01

    Full Text Available The identification of mycobacteria is essential because tuberculosis (TB and mycobacteriosis are clinically indistinguishable and require different therapeutic regimens. The traditional phenotypic method is time consuming and may last up to 60 days. Indeed, rapid, affordable, specific and easy-to-perform identification methods are needed. We have previously described a polymerase chain reaction-based method called a mycobacteria mobility shift assay (MMSA that was designed for Mycobacterium tuberculosis complex (MTC and nontuberculous mycobacteria (NTM species identification. The aim of this study was to assess the MMSA for the identification of MTC and NTM clinical isolates and to compare its performance with that of the PRA-hsp65 method. A total of 204 clinical isolates (102 NTM and 102 MTC were identified by the MMSA and PRA-hsp65. For isolates for which these methods gave discordant results, definitive species identification was obtained by sequencing fragments of the 16S rRNA and hsp65 genes. Both methods correctly identified all MTC isolates. Among the NTM isolates, the MMSA alone assigned 94 (92.2% to a complex or species, whereas the PRA-hsp65 method assigned 100% to a species. A 91.5% agreement was observed for the 94 NTM isolates identified by both methods. The MMSA provided correct identification for 96.8% of the NTM isolates compared with 94.7% for PRA-hsp65. The MMSA is a suitable auxiliary method for routine use for the rapid identification of mycobacteria.

  20. Mycobacteria mobility shift assay: a method for the rapid identification of Mycobacterium tuberculosis and nontuberculous mycobacteria.

    Science.gov (United States)

    Wildner, Letícia Muraro; Bazzo, Maria Luiza; Liedke, Susie Coutinho; Nogueira, Christiane Lourenço; Segat, Gabriela; Senna, Simone Gonçalves; Schlindwein, Aline Daiane; Oliveira, Jaquelline Germano de; Rovaris, Darcita B; Bonjardim, Claudio A; Kroon, Erna G; Ferreira, Paulo C P

    2014-06-01

    The identification of mycobacteria is essential because tuberculosis (TB) and mycobacteriosis are clinically indistinguishable and require different therapeutic regimens. The traditional phenotypic method is time consuming and may last up to 60 days. Indeed, rapid, affordable, specific and easy-to-perform identification methods are needed. We have previously described a polymerase chain reaction-based method called a mycobacteria mobility shift assay (MMSA) that was designed for Mycobacterium tuberculosis complex (MTC) and nontuberculous mycobacteria (NTM) species identification. The aim of this study was to assess the MMSA for the identification of MTC and NTM clinical isolates and to compare its performance with that of the PRA-hsp65 method. A total of 204 clinical isolates (102 NTM and 102 MTC) were identified by the MMSA and PRA-hsp65. For isolates for which these methods gave discordant results, definitive species identification was obtained by sequencing fragments of the 16S rRNA and hsp65 genes. Both methods correctly identified all MTC isolates. Among the NTM isolates, the MMSA alone assigned 94 (92.2%) to a complex or species, whereas the PRA-hsp65 method assigned 100% to a species. A 91.5% agreement was observed for the 94 NTM isolates identified by both methods. The MMSA provided correct identification for 96.8% of the NTM isolates compared with 94.7% for PRA-hsp65. The MMSA is a suitable auxiliary method for routine use for the rapid identification of mycobacteria.

  1. Rapid identification of Stenotrophomonas maltophilia by peptide nucleic acid fluorescence in situ hybridization

    DEFF Research Database (Denmark)

    Knudsen, Nanna Reumert; Rasmussen, A. K. I.; Fiandaca, M. J.

    2014-01-01

    The objective of this study was to develop a novel peptide nucleic acid (PNA) probe for Stenotrophomonas maltophilia identification by fluorescence in situ hybridization (FISH). The probe was evaluated using 33 human and veterinary clinical S. maltophilia isolates and 45 reference strains...... horses. Altogether the study shows that this species-specific PNA FISH probe facilitates rapid detection of S. maltophilia in biological specimens....

  2. Rapid Identification and Characterization of Formulated Protein Products by Raman Spectroscopy Coupled with Discriminant Analysis.

    Science.gov (United States)

    Cao, Xiaolin; Zhou, Dan; Loussaert, James A; Meriage, David S; Levine, Joseph D; Gabrielson, John P; Wen, Zai-Qing

    2016-01-01

    The rapid identification of protein drug products for packaging and receiving can significantly reduce disposition cycle time, and thereby improve the efficiency and productivity of the supply chain to better meet the needs of patients. In this feasibility study, we demonstrate a novel methodology that combines Raman spectroscopy with discriminant analysis that can be used for rapid identification or verification of finished products. With this methodology, Raman spectra of formulated therapeutic proteins were collected non-invasively with the samples either in a quartz cuvette or in the original glass vials, and analyzed without subtraction of buffer or placebo solutions. The algorithm used for the discriminant analysis was Mahalanobis distance by principal component analysis with residuals. In addition to product identification, the methodology has the potential to be used for characterizing formulated proteins when exposed to external stresses based on the changes of Mahalanobis distances. The rapid identification of protein drug products for packaging and receiving can significantly reduce disposition cycle time, and thereby improve the efficiency and productivity of the supply chain. In this study, we demonstrate a novel methodology that combines Raman spectroscopy with discriminant analysis to rapidly identify formulated proteins non-invasively. © PDA, Inc. 2016.

  3. Comparison between MALDI-TOF MS and FilmArray Blood Culture Identification panel for rapid identification of yeast from positive blood culture.

    Science.gov (United States)

    Paolucci, M; Foschi, C; Tamburini, M V; Ambretti, S; Lazzarotto, T; Landini, M P

    2014-09-01

    In this study we evaluated MALDI-TOF MS and FilmArray methods for the rapid identification of yeast from positive blood cultures. FilmArray correctly identified 20/22 of yeast species, while MALDI-TOF MS identified 9/22. FilmArray is a reliable and rapid identification system for the direct identification of yeasts from positive blood cultures. Copyright © 2014 Elsevier B.V. All rights reserved.

  4. Pyrosequencing as a tool for rapid fish species identification and commercial fraud detection.

    Science.gov (United States)

    De Battisti, Cristian; Marciano, Sabrina; Magnabosco, Cristian; Busato, Sara; Arcangeli, Giuseppe; Cattoli, Giovanni

    2014-01-08

    The increased consumption of fish products, as well as the occurrence of exotic fish species in the Mediterranean Sea and in the fish market, has increased the risk of commercial fraud. Furthermore, the great amount of processed seafood products has greatly limited the application of classic identification systems. DNA-based identification allows a clear and unambiguous detection of polymorphisms between species, permitting differentiation and identification of both commercial fraud and introduction of species with potential toxic effects on humans. In this study, a novel DNA-based approach for differentiation of fish species based on pyrosequencing technology has been developed. Raw and processed fish products were tested, and up to 25 species of fish belonging to Clupeiformes and Pleuronectiformes groups were uniquely and rapidly identified. The proper identification based on short and unique genetic sequence signatures demonstrates that this approach is promising and cost-effective for large-scale surveys.

  5. Dynamical recurrent neuro-fuzzy identification schemes employing switching parameter hopping.

    Science.gov (United States)

    Theodoridis, Dimitrios; Boutalis, Yiannis; Christodoulou, Manolis

    2012-04-01

    In this paper we analyze the identification problem which consists of choosing an appropriate identification model and adjusting its parameters according to some adaptive law, such that the response of the model to an input signal (or a class of input signals), approximates the response of the real system for the same input. For identification models we use fuzzy-recurrent high order neural networks. High order networks are expansions of the first-order Hopfield and Cohen-Grossberg models that allow higher order interactions between neurons. The underlying fuzzy model is of Mamdani type assuming a standard defuzzification procedure such as the weighted average. Learning laws are proposed which ensure that the identification error converges to zero exponentially fast or to a residual set when a modeling error is applied. There are two core ideas in the proposed method: (1) Several high order neural networks are specialized to work around fuzzy centers, separating in this way the system into neuro-fuzzy subsystems, and (2) the use of a novel method called switching parameter hopping against the commonly used projection in order to restrict the weights and avoid drifting to infinity.

  6. Rapid identification of Yersinia pestis and Brucella melitensis by chip-based continuous flow PCR

    Science.gov (United States)

    Dietzsch, Michael; Hlawatsch, Nadine; Melzer, Falk; Tomaso, Herbert; Gärtner, Claudia; Neubauer, Heinrich

    2012-06-01

    To combat the threat of biological agents like Yersinia pestis and Brucella melitensis in bioterroristic scenarios requires fast, easy-to-use and safe identification systems. In this study we describe a system for rapid amplification of specific genetic markers for the identification of Yersinia pestis and Brucella melitensis. Using chip based PCR and continuous flow technology we were able to amplify the targets simultaneously with a 2-step reaction profile within 20 minutes. The subsequent analysis of amplified fragments by standard gel electrophoresis requires another 45 minutes. We were able to detect both pathogens within 75 minutes being much faster than most other nucleic acid amplification technologies.

  7. Rapid identification of sequences for orphan enzymes to power accurate protein annotation.

    Directory of Open Access Journals (Sweden)

    Kevin R Ramkissoon

    Full Text Available The power of genome sequencing depends on the ability to understand what those genes and their proteins products actually do. The automated methods used to assign functions to putative proteins in newly sequenced organisms are limited by the size of our library of proteins with both known function and sequence. Unfortunately this library grows slowly, lagging well behind the rapid increase in novel protein sequences produced by modern genome sequencing methods. One potential source for rapidly expanding this functional library is the "back catalog" of enzymology--"orphan enzymes," those enzymes that have been characterized and yet lack any associated sequence. There are hundreds of orphan enzymes in the Enzyme Commission (EC database alone. In this study, we demonstrate how this orphan enzyme "back catalog" is a fertile source for rapidly advancing the state of protein annotation. Starting from three orphan enzyme samples, we applied mass-spectrometry based analysis and computational methods (including sequence similarity networks, sequence and structural alignments, and operon context analysis to rapidly identify the specific sequence for each orphan while avoiding the most time- and labor-intensive aspects of typical sequence identifications. We then used these three new sequences to more accurately predict the catalytic function of 385 previously uncharacterized or misannotated proteins. We expect that this kind of rapid sequence identification could be efficiently applied on a larger scale to make enzymology's "back catalog" another powerful tool to drive accurate genome annotation.

  8. Rapid Identification of Sequences for Orphan Enzymes to Power Accurate Protein Annotation

    Science.gov (United States)

    Ojha, Sunil; Watson, Douglas S.; Bomar, Martha G.; Galande, Amit K.; Shearer, Alexander G.

    2013-01-01

    The power of genome sequencing depends on the ability to understand what those genes and their proteins products actually do. The automated methods used to assign functions to putative proteins in newly sequenced organisms are limited by the size of our library of proteins with both known function and sequence. Unfortunately this library grows slowly, lagging well behind the rapid increase in novel protein sequences produced by modern genome sequencing methods. One potential source for rapidly expanding this functional library is the “back catalog” of enzymology – “orphan enzymes,” those enzymes that have been characterized and yet lack any associated sequence. There are hundreds of orphan enzymes in the Enzyme Commission (EC) database alone. In this study, we demonstrate how this orphan enzyme “back catalog” is a fertile source for rapidly advancing the state of protein annotation. Starting from three orphan enzyme samples, we applied mass-spectrometry based analysis and computational methods (including sequence similarity networks, sequence and structural alignments, and operon context analysis) to rapidly identify the specific sequence for each orphan while avoiding the most time- and labor-intensive aspects of typical sequence identifications. We then used these three new sequences to more accurately predict the catalytic function of 385 previously uncharacterized or misannotated proteins. We expect that this kind of rapid sequence identification could be efficiently applied on a larger scale to make enzymology’s “back catalog” another powerful tool to drive accurate genome annotation. PMID:24386392

  9. Some Numerical Methods for Exponential Analysis with Connection to a General Identification Scheme for Linear Processes

    Science.gov (United States)

    1980-11-01

    identify by block number) System- identification, Prony , exponential analysis, Singularity Expansion Method, transient 20 BTAT(Continue o...produces a "best fit " to the wave- form under analysis in the mean-square 3ense, that is, it minimizes the mean-square error. Some other methods that have...herein to describe the Prony procedure (24], the so-called least squares Prony procedure [9], and the pencil-of-functions method of Jain [191. The

  10. A Pre-Detection Based Anti-Collision Algorithm with Adjustable Slot Size Scheme for Tag Identification

    Directory of Open Access Journals (Sweden)

    Chiu-Kuo LIANG

    2015-06-01

    Full Text Available One of the research areas in RFID systems is a tag anti-collision protocol; how to reduce identification time with a given number of tags in the field of an RFID reader. There are two types of tag anti-collision protocols for RFID systems: tree based algorithms and slotted aloha based algorithms. Many anti-collision algorithms have been proposed in recent years, especially in tree based protocols. However, there still have challenges on enhancing the system throughput and stability due to the underlying technologies had faced different limitation in system performance when network density is high. Particularly, the tree based protocols had faced the long identification delay. Recently, a Hybrid Hyper Query Tree (H2QT protocol, which is a tree based approach, was proposed and aiming to speedup tag identification in large scale RFID systems. The main idea of H2QT is to track the tag response and try to predict the distribution of tag IDs in order to reduce collisions. In this paper, we propose a pre-detection tree based algorithm, called the Adaptive Pre-Detection Broadcasting Query Tree algorithm (APDBQT, to avoid those unnecessary queries. Our proposed APDBQT protocol can reduce not only the collisions but the idle cycles as well by using pre-detection scheme and adjustable slot size mechanism. The simulation results show that our proposed technique provides superior performance in high density environments. It is shown that the APDBQT is effective in terms of increasing system throughput and minimizing identification delay.

  11. A rapid and direct real time PCR-based method for identification of Salmonella spp

    DEFF Research Database (Denmark)

    Rodriguez-Lazaro, D.; Hernández, Marta; Esteve, T.

    2003-01-01

    The aim of this work was the validation of a rapid, real-time PCR assay based on TaqMan((R)) technology for the unequivocal identification of Salmonella spp. to be used directly on an agar-grown colony. A real-time PCR system targeting at the Salmonella spp. invA gene was optimized and validated...... to be especially convenient because the pre-mix containing all PCR reagents except for the bacterial cells could be kept at -20 degreesC for at least I month before its use. The optimized TaqMan((R)) real-time PCR assay is a useful, simple and rapid method for routine identification of Salmonella spp...

  12. Rapid Identification of Ralstonia solanacearum by the Direct Colony TLC and Simple TLC

    OpenAIRE

    Rahman, Md Zahidur; Mian, Ismail Hossain; Khan, Abu Ashraf; Furuya, Naruto; Matsuyama, Nobuaki

    1998-01-01

    Rapid identification of bacterial wilt pathogen, Ralstonia solanacearum was conducted by the direct colony TLC and simple TLC methods. All fourteen isolates from different host plants in Bangladesh exhibited similar chromatograms with R. solanacearum ATCC 11696 the type strain by the direct colony TLC method. This result indicated that all the isolates were R. solanacearum. Physiological and biochemical tests on the fourteen isolates also verified that they were R. solanacearum. Seven strains...

  13. Rapid Identification of Black Grain Eumycetoma Causative Agents Using Rolling Circle Amplification

    Science.gov (United States)

    Ahmed, Sarah A.; van den Ende, Bert H. G. Gerrits; Fahal, Ahmed H.; van de Sande, Wendy W. J.; de Hoog, G. S.

    2014-01-01

    Accurate identification of mycetoma causative agent is a priority for treatment. However, current identification tools are far from being satisfactory for both reliable diagnosis and epidemiological investigations. A rapid, simple, and highly efficient molecular based method for identification of agents of black grain eumycetoma is introduced, aiming to improve diagnostic in endemic areas. Rolling Circle Amplification (RCA) uses species-specific padlock probes and isothermal DNA amplification. The tests were based on ITS sequences and developed for Falciformispora senegalensis, F. tompkinsii, Madurella fahalii, M. mycetomatis, M. pseudomycetomatis, M. tropicana, Medicopsis romeroi, and Trematosphaeria grisea. With the isothermal RCA assay, 62 isolates were successfully identified with 100% specificity and no cross reactivity or false results. The main advantage of this technique is the low-cost, high specificity, and simplicity. In addition, it is highly reproducible and can be performed within a single day. PMID:25474355

  14. Rapid identification of black grain eumycetoma causative agents using rolling circle amplification.

    Directory of Open Access Journals (Sweden)

    Sarah A Ahmed

    2014-12-01

    Full Text Available Accurate identification of mycetoma causative agent is a priority for treatment. However, current identification tools are far from being satisfactory for both reliable diagnosis and epidemiological investigations. A rapid, simple, and highly efficient molecular based method for identification of agents of black grain eumycetoma is introduced, aiming to improve diagnostic in endemic areas. Rolling Circle Amplification (RCA uses species-specific padlock probes and isothermal DNA amplification. The tests were based on ITS sequences and developed for Falciformispora senegalensis, F. tompkinsii, Madurella fahalii, M. mycetomatis, M. pseudomycetomatis, M. tropicana, Medicopsis romeroi, and Trematosphaeria grisea. With the isothermal RCA assay, 62 isolates were successfully identified with 100% specificity and no cross reactivity or false results. The main advantage of this technique is the low-cost, high specificity, and simplicity. In addition, it is highly reproducible and can be performed within a single day.

  15. A PSO Driven Intelligent Model Updating and Parameter Identification Scheme for Cable-Damper System

    Directory of Open Access Journals (Sweden)

    Danhui Dan

    2015-01-01

    Full Text Available The precise measurement of the cable force is very important for monitoring and evaluating the operation status of cable structures such as cable-stayed bridges. The cable system should be installed with lateral dampers to reduce the vibration, which affects the precise measurement of the cable force and other cable parameters. This paper suggests a cable model updating calculation scheme driven by the particle swarm optimization (PSO algorithm. By establishing a finite element model considering the static geometric nonlinearity and stress-stiffening effect firstly, an automatically finite element method model updating powered by PSO algorithm is proposed, with the aims to identify the cable force and relevant parameters of cable-damper system precisely. Both numerical case studies and full-scale cable tests indicated that, after two rounds of updating process, the algorithm can accurately identify the cable force, moment of inertia, and damping coefficient of the cable-damper system.

  16. MALDI-TOF mass spectrometry for the rapid identification of aetiological agents of sepsis

    Directory of Open Access Journals (Sweden)

    Roberto Degl’Innocenti

    2013-04-01

    Full Text Available Introduction: The MALDI-TOF has recently become part of the methods of microbiological investigation in many laboratories of bacteriology with advantages both practical and economical.The use of this technique for the rapid identification of the causative agents of sepsis is of strategic importance to the ability to provide the clinician with useful information for a prompt and rapid establishment of an empirical antimicrobial “targeted” therapy. Methods: It was tested a total of 343 positive blood culture bottles from 211 patients. The samples after collection were incubated in the BACTEC FX (Becton Dickinson, USA. From these bottles were taken a few milliliters of broth culture and transferred into a vacutainer tube containing gel. This was centrifuged, the supernatant was decanted, and finally recovered the bacterial suspension on the gel. With micro-organisms recovered in this way, after several washes with distilled water, was prepared a slide for microscopic examination with Gram stain, and a plate for mass spectrometry (MS-Vitek, bioMérieux, France.Then, the same samples were inoculated on solid agar media according to the protocol in use in our laboratory.The next day was checked the possible bacterial growth on solid media; we then proceeded to the identification of the colonies by Vitek MS and / or with the system Vitek2 (bioMérieux, France. Results: 258 (75.2% positive vials show concordant results between direct identification and identification after growth on agar. For 83 (24.2% positive bottles there has been full compliance with the microscopic examination but not with culture. In particular, two bottles (0.6% have given complete discordance between the direct identification and that after growth. Conclusions: The protocol we use for the direct identification of organisms responsible for sepsis, directly on positive bottles, seems to be a quick and inexpensive procedure, which in less than 60 minutes can give valuable

  17. An automatic microseismic or acoustic emission arrival identification scheme with deep recurrent neural networks

    Science.gov (United States)

    Zheng, Jing; Lu, Jiren; Peng, Suping; Jiang, Tianqi

    2018-02-01

    The conventional arrival pick-up algorithms cannot avoid the manual modification of the parameters for the simultaneous identification of multiple events under different signal-to-noise ratios (SNRs). Therefore, in order to automatically obtain the arrivals of multiple events with high precision under different SNRs, in this study an algorithm was proposed which had the ability to pick up the arrival of microseismic or acoustic emission events based on deep recurrent neural networks. The arrival identification was performed using two important steps, which included a training phase and a testing phase. The training process was mathematically modelled by deep recurrent neural networks using Long Short-Term Memory architecture. During the testing phase, the learned weights were utilized to identify the arrivals through the microseismic/acoustic emission data sets. The data sets were obtained by rock physics experiments of the acoustic emission. In order to obtain the data sets under different SNRs, this study added random noise to the raw experiments' data sets. The results showed that the outcome of the proposed method was able to attain an above 80 per cent hit-rate at SNR 0 dB, and an approximately 70 per cent hit-rate at SNR -5 dB, with an absolute error in 10 sampling points. These results indicated that the proposed method had high selection precision and robustness.

  18. Spatial-Temporal Feature Analysis on Single-Trial Event Related Potential for Rapid Face Identification.

    Science.gov (United States)

    Jiang, Lei; Wang, Yun; Cai, Bangyu; Wang, Yueming; Wang, Yiwen

    2017-01-01

    The event-related potential (ERP) is the brain response measured in electroencephalography (EEG), which reflects the process of human cognitive activity. ERP has been introduced into brain computer interfaces (BCIs) to communicate the computer with the subject's intention. Due to the low signal-to-noise ratio of EEG, most ERP studies are based on grand-averaging over many trials. Recently single-trial ERP detection attracts more attention, which enables real time processing tasks as rapid face identification. All the targets needed to be retrieved may appear only once, and there is no knowledge of target label for averaging. More interestingly, how the features contribute temporally and spatially to single-trial ERP detection has not been fully investigated. In this paper, we propose to implement a local-learning-based (LLB) feature extraction method to investigate the importance of spatial-temporal components of ERP in a task of rapid face identification using single-trial detection. Comparing to previous methods, LLB method preserves the nonlinear structure of EEG signal distribution, and analyze the importance of original spatial-temporal components via optimization in feature space. As a data-driven methods, the weighting of the spatial-temporal component does not depend on the ERP detection method. The importance weights are optimized by making the targets more different from non-targets in feature space, and regularization penalty is introduced in optimization for sparse weights. This spatial-temporal feature extraction method is evaluated on the EEG data of 15 participants in performing a face identification task using rapid serial visual presentation paradigm. Comparing with other methods, the proposed spatial-temporal analysis method uses sparser (only 10% of the total) features, and could achieve comparable performance (98%) of single-trial ERP detection as the whole features across different detection methods. The interesting finding is that the N250 is

  19. A Very Low-Speed Sensorless Induction Motor Drive with Online Stator Resistance identification scheme

    Directory of Open Access Journals (Sweden)

    M. Zaky

    2008-03-01

    Full Text Available Recently, speed sensorless control of induction motor drives received great attention to avoid the different problems associated with direct speed sensors. However, low speed operation with robustness against parameter variations remains an area of research for sensorless systems. Stator resistance is of greatest importance for good operation of speed sensorless systems in low speed region. In this paper, a sliding mode current observer for an induction motor is presented. An estimation algorithm based on this observer in conjunction with Popov's hyper-stability theory is proposed to calculate the speed and stator resistance independently. The proposed speed observer with parallel stator resistance identification is firstly verified by simulation. Experimental results are included as well to demonstrate the good performance of the proposed observer and estimation algorithms at low speed.

  20. Stability improvement of induction motor based on stator parameter identification scheme

    Science.gov (United States)

    Kang, Shanlin; Kang, Yuzhe; Chen, Jingwei

    2008-10-01

    A new method for stator resistance identification based on wavelet network is presented to improve the operating performance of induction motor. The ripple of the torque is kept constant, resulting in a variable switching frequency, which depends on the tolerance band of the control. Due to wavelet transform behaving good localization property in both time and frequency space and multi-scale property, the wavelet function is adopted as the basic function of neural network. The current error and the change in the current error are the inputs of the wavelet network and the stator resistance error is the output of the wavelet network. The network parameters are initialized by the improved least squares algorithm. The parameters of wavelet network are tuned online to approximate the unknown nonlinear model with an appropriately chosen adaptive mechanism. The proposed method is proved to be efficient to reduce the torque ripple and current ripple by detailed comparison simulation results.

  1. Rapid identification of oral Actinomyces species cultivated from subgingival biofilm by MALDI-TOF-MS

    Science.gov (United States)

    Stingu, Catalina S.; Borgmann, Toralf; Rodloff, Arne C.; Vielkind, Paul; Jentsch, Holger; Schellenberger, Wolfgang; Eschrich, Klaus

    2015-01-01

    Background Actinomyces are a common part of the residential flora of the human intestinal tract, genitourinary system and skin. Isolation and identification of Actinomyces by conventional methods is often difficult and time consuming. In recent years, matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF-MS) has become a rapid and simple method to identify bacteria. Objective The present study evaluated a new in-house algorithm using MALDI-TOF-MS for rapid identification of different species of oral Actinomyces cultivated from subgingival biofilm. Design Eleven reference strains and 674 clinical strains were used in this study. All the strains were preliminarily identified using biochemical methods and then subjected to MALDI-TOF-MS analysis using both similarity-based analysis and classification methods (support vector machine [SVM]). The genotype of the reference strains and of 232 clinical strains was identified by sequence analysis of the 16S ribosomal RNA (rRNA). Results The sequence analysis of the 16S rRNA gene of all references strains confirmed their previous identification. The MALDI-TOF-MS spectra obtained from the reference strains and the other clinical strains undoubtedly identified as Actinomyces by 16S rRNA sequencing were used to create the mass spectra reference database. Already a visual inspection of the mass spectra of different species reveals both similarities and differences. However, the differences between them are not large enough to allow a reliable differentiation by similarity analysis. Therefore, classification methods were applied as an alternative approach for differentiation and identification of Actinomyces at the species level. A cross-validation of the reference database representing 14 Actinomyces species yielded correct results for all species which were represented by more than two strains in the database. Conclusions Our results suggest that a combination of MALDI-TOF-MS with powerful

  2. Rapid identification of oral Actinomyces species cultivated from subgingival biofilm by MALDI-TOF-MS

    Directory of Open Access Journals (Sweden)

    Catalina S. Stingu

    2015-01-01

    Full Text Available Background: Actinomyces are a common part of the residential flora of the human intestinal tract, genitourinary system and skin. Isolation and identification of Actinomyces by conventional methods is often difficult and time consuming. In recent years, matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF-MS has become a rapid and simple method to identify bacteria. Objective: The present study evaluated a new in-house algorithm using MALDI-TOF-MS for rapid identification of different species of oral Actinomyces cultivated from subgingival biofilm. Design: Eleven reference strains and 674 clinical strains were used in this study. All the strains were preliminarily identified using biochemical methods and then subjected to MALDI-TOF-MS analysis using both similarity-based analysis and classification methods (support vector machine [SVM]. The genotype of the reference strains and of 232 clinical strains was identified by sequence analysis of the 16S ribosomal RNA (rRNA. Results: The sequence analysis of the 16S rRNA gene of all references strains confirmed their previous identification. The MALDI-TOF-MS spectra obtained from the reference strains and the other clinical strains undoubtedly identified as Actinomyces by 16S rRNA sequencing were used to create the mass spectra reference database. Already a visual inspection of the mass spectra of different species reveals both similarities and differences. However, the differences between them are not large enough to allow a reliable differentiation by similarity analysis. Therefore, classification methods were applied as an alternative approach for differentiation and identification of Actinomyces at the species level. A cross-validation of the reference database representing 14 Actinomyces species yielded correct results for all species which were represented by more than two strains in the database. Conclusions: Our results suggest that a combination of MALDI

  3. Toward Automating HIV Identification: Machine Learning for Rapid Identification of HIV-Related Social Media Data.

    Science.gov (United States)

    Young, Sean D; Yu, Wenchao; Wang, Wei

    2017-02-01

    "Social big data" from technologies such as social media, wearable devices, and online searches continue to grow and can be used as tools for HIV research. Although researchers can uncover patterns and insights associated with HIV trends and transmission, the review process is time consuming and resource intensive. Machine learning methods derived from computer science might be used to assist HIV domain experts by learning how to rapidly and accurately identify patterns associated with HIV from a large set of social data. Using an existing social media data set that was associated with HIV and coded by an HIV domain expert, we tested whether 4 commonly used machine learning methods could learn the patterns associated with HIV risk behavior. We used the 10-fold cross-validation method to examine the speed and accuracy of these models in applying that knowledge to detect HIV content in social media data. Logistic regression and random forest resulted in the highest accuracy in detecting HIV-related social data (85.3%), whereas the Ridge Regression Classifier resulted in the lowest accuracy. Logistic regression yielded the fastest processing time (16.98 seconds). Machine learning can enable social big data to become a new and important tool in HIV research, helping to create a new field of "digital HIV epidemiology." If a domain expert can identify patterns in social data associated with HIV risk or HIV transmission, machine learning models could quickly and accurately learn those associations and identify potential HIV patterns in large social data sets.

  4. Rapid Multi-Damage Identification for Health Monitoring of Laminated Composites Using Piezoelectric Wafer Sensor Arrays

    Science.gov (United States)

    Si, Liang; Wang, Qian

    2016-01-01

    Through the use of the wave reflection from any damage in a structure, a Hilbert spectral analysis-based rapid multi-damage identification (HSA-RMDI) technique with piezoelectric wafer sensor arrays (PWSA) is developed to monitor and identify the presence, location and severity of damage in carbon fiber composite structures. The capability of the rapid multi-damage identification technique to extract and estimate hidden significant information from the collected data and to provide a high-resolution energy-time spectrum can be employed to successfully interpret the Lamb waves interactions with single/multiple damage. Nevertheless, to accomplish the precise positioning and effective quantification of multiple damage in a composite structure, two functional metrics from the RMDI technique are proposed and used in damage identification, which are the energy density metric and the energy time-phase shift metric. In the designed damage experimental tests, invisible damage to the naked eyes, especially delaminations, were detected in the leftward propagating waves as well as in the selected sensor responses, where the time-phase shift spectra could locate the multiple damage whereas the energy density spectra were used to quantify the multiple damage. The increasing damage was shown to follow a linear trend calculated by the RMDI technique. All damage cases considered showed completely the developed RMDI technique potential as an effective online damage inspection and assessment tool. PMID:27153070

  5. Rapid Multi-Damage Identification for Health Monitoring of Laminated Composites Using Piezoelectric Wafer Sensor Arrays.

    Science.gov (United States)

    Si, Liang; Wang, Qian

    2016-05-04

    Through the use of the wave reflection from any damage in a structure, a Hilbert spectral analysis-based rapid multi-damage identification (HSA-RMDI) technique with piezoelectric wafer sensor arrays (PWSA) is developed to monitor and identify the presence, location and severity of damage in carbon fiber composite structures. The capability of the rapid multi-damage identification technique to extract and estimate hidden significant information from the collected data and to provide a high-resolution energy-time spectrum can be employed to successfully interpret the Lamb waves interactions with single/multiple damage. Nevertheless, to accomplish the precise positioning and effective quantification of multiple damage in a composite structure, two functional metrics from the RMDI technique are proposed and used in damage identification, which are the energy density metric and the energy time-phase shift metric. In the designed damage experimental tests, invisible damage to the naked eyes, especially delaminations, were detected in the leftward propagating waves as well as in the selected sensor responses, where the time-phase shift spectra could locate the multiple damage whereas the energy density spectra were used to quantify the multiple damage. The increasing damage was shown to follow a linear trend calculated by the RMDI technique. All damage cases considered showed completely the developed RMDI technique potential as an effective online damage inspection and assessment tool.

  6. Rapid Multi-Damage Identification for Health Monitoring of Laminated Composites Using Piezoelectric Wafer Sensor Arrays

    Directory of Open Access Journals (Sweden)

    Liang Si

    2016-05-01

    Full Text Available Through the use of the wave reflection from any damage in a structure, a Hilbert spectral analysis-based rapid multi-damage identification (HSA-RMDI technique with piezoelectric wafer sensor arrays (PWSA is developed to monitor and identify the presence, location and severity of damage in carbon fiber composite structures. The capability of the rapid multi-damage identification technique to extract and estimate hidden significant information from the collected data and to provide a high-resolution energy-time spectrum can be employed to successfully interpret the Lamb waves interactions with single/multiple damage. Nevertheless, to accomplish the precise positioning and effective quantification of multiple damage in a composite structure, two functional metrics from the RMDI technique are proposed and used in damage identification, which are the energy density metric and the energy time-phase shift metric. In the designed damage experimental tests, invisible damage to the naked eyes, especially delaminations, were detected in the leftward propagating waves as well as in the selected sensor responses, where the time-phase shift spectra could locate the multiple damage whereas the energy density spectra were used to quantify the multiple damage. The increasing damage was shown to follow a linear trend calculated by the RMDI technique. All damage cases considered showed completely the developed RMDI technique potential as an effective online damage inspection and assessment tool.

  7. Rapid and accurate identification of microorganisms contaminating cosmetic products based on DNA sequence homology.

    Science.gov (United States)

    Fujita, Y; Shibayama, H; Suzuki, Y; Karita, S; Takamatsu, S

    2005-12-01

    The aim of this study was to develop rapid and accurate procedures to identify microorganisms contaminating cosmetic products, based on the identity of the nucleotide sequences of the internal transcribed spacer (ITS) region of the ribosomal RNA coding DNA (rDNA). Five types of microorganisms were isolated from the inner portion of lotion bottle caps, skin care lotions, and cleansing gels. The rDNA ITS region of microorganisms was amplified through the use of colony-direct PCR or ordinal PCR using DNA extracts as templates. The nucleotide sequences of the amplified DNA were determined and subjected to homology search of a publicly available DNA database. Thereby, we obtained DNA sequences possessing high similarity with the query sequences from the databases of all the five organisms analyzed. The traditional identification procedure requires expert skills, and a time period of approximately 1 month to identify the microorganisms. On the contrary, 3-7 days were sufficient to complete all the procedures employed in the current method, including isolation and cultivation of organisms, DNA sequencing, and the database homology search. Moreover, it was possible to develop the skills necessary to perform the molecular techniques required for the identification procedures within 1 week. Consequently, the current method is useful for rapid and accurate identification of microorganisms, contaminating cosmetics.

  8. Rapid intrinsic fluorescence method for direct identification of pathogens in blood cultures.

    Science.gov (United States)

    Walsh, John D; Hyman, Jay M; Borzhemskaya, Larisa; Bowen, Ann; McKellar, Caroline; Ullery, Michael; Mathias, Erin; Ronsick, Christopher; Link, John; Wilson, Mark; Clay, Bradford; Robinson, Ron; Thorpe, Thurman; van Belkum, Alex; Dunne, W Michael

    2013-11-19

    A positive blood culture is a critical result that requires prompt identification of the causative agent. This article describes a simple method to identify microorganisms from positive blood culture broth within the time taken to perform a Gram stain (identification of the etiologic agent may benefit the clinical management of sepsis. Further evaluation is now warranted to determine the performance of the method using clinical blood culture specimens. Physicians often require the identity of the infective agent in order to make life-saving adjustments to empirical therapy or to switch to less expensive and/or more targeted antimicrobials. However, standard identification procedures take up to 2 days after a blood culture is signaled positive, and even most rapid molecular techniques take several hours to provide a result. Other techniques are faster (e.g., matrix-assisted laser desorption ionization-time of flight [MALDI-TOF] mass spectrometry) but require time-consuming manual processing steps and expensive equipment. There remains a clear need for a simple, inexpensive method to rapidly identify microorganisms directly from positive blood cultures. The promising new method described in this research article can identify microorganisms in minutes by optical spectroscopy, thus permitting the lab to simultaneously report the presence of a positive blood culture and the organism's identity.

  9. An Ultrafast Maximum Power Point Setting Scheme for Photovoltaic Arrays Using Model Parameter Identification

    Directory of Open Access Journals (Sweden)

    Zhaohui Cen

    2015-01-01

    Full Text Available Maximum power point tracking (MPPT for photovoltaic (PV arrays is essential to optimize conversion efficiency under variable and nonuniform irradiance conditions. Unfortunately, conventional MPPT algorithms such as perturb and observe (P&O, incremental conductance, and current sweep method need to iterate command current or voltage and frequently operate power converters with associated losses. Under partial overcast conditions, tracking the real MPP in multipeak P-I or P-V curve model becomes highly challenging, with associated increase in search time and converter operation, leading to unnecessary power being lost in the MPP tracking process. In this paper, the noted drawbacks in MPPT-controlled converters are addressed. In order to separate the search algorithms from converter operation, a model parameter identification approach is presented to estimate insolation conditions of each PV panel and build a real-time overall P-I curve of PV arrays. Subsequently a simple but effective global MPPT algorithm is proposed to track the MPP in the overall P-I curve obtained from the identified PV array model, ensuring that the converter works at the MPP. The novel MPPT is ultrafast, resulting in conserved power in the tracking process. Finally, simulations in different scenarios are executed to validate the novel scheme’s effectiveness and advantages.

  10. Prospective study of the performance of vibrational spectroscopies for rapid identification of bacterial and fungal pathogens recovered from blood cultures

    NARCIS (Netherlands)

    K. Maquelin (Kees); C. Kirschner; L.P. Choo-Smith; N.A. Ngo-Thi; T. van Vreeswijk; M. Stammler; H.P. Endtz (Hubert); H.A. Bruining (Hajo); D. Naumann; G.J. Puppels (Gerwin)

    2003-01-01

    textabstractRapid identification of microbial pathogens reduces infection-related morbidity and mortality of hospitalized patients. Raman spectra and Fourier transform infrared (IR) spectra constitute highly specific spectroscopic fingerprints of microorganisms by which they can

  11. Etiological Analysis of Fungal Keratitis and Rapid Identification of Predominant Fungal Pathogens.

    Science.gov (United States)

    He, Dan; Hao, Jilong; Gao, Song; Wan, Xue; Wang, Wanting; Shan, Qiushi; Wang, Li

    2016-02-01

    Fungal keratitis is a worldwide-distributed refractory and potentially blinding ocular infection caused by various fungi. It is necessary to investigate the etiological and epidemiological characteristics of this disease and establish a rapid and specific pathogenic identification method. Here, we isolated and identified fungal pathogens of 275 patients with presumed fungal keratitis from Jilin Province, China, and conducted statistical analyses of epidemiological information. The positive rate of fungal culture was 72.0 %. Fusarium sp. was the most common genus among 210 fungal isolates. The predominant species were Fusarium solani, Aspergillus fumigatus, and Candida glabrata, which accounted for over 50 % of the isolated organisms. Corneal trauma and previous use of drugs were the most important predisposing factors. In addition, a multiplex polymerase chain reaction (PCR) was designed with species-specific primers of the three species that could identify them with amplicons of approximately 330 bp from F. solani, 275 bp from A. fumigatus, and 230 bp from C. glabrata. Additionally, PCR with fungal universal primers and multiplex PCR were performed using DNA prepared by an improved DNA extraction method from corneal scrapings. With this method, fungal pathogens from corneal scrapings could be specifically and rapidly identified within 8 h. The culture-independent rapid identification of corneal scrapings may have great significance for the early diagnosis and treatment of fungal keratitis.

  12. Evaluation of Verigene Blood Culture Test Systems for Rapid Identification of Positive Blood Cultures

    Directory of Open Access Journals (Sweden)

    Jae-Seok Kim

    2016-01-01

    Full Text Available The performance of molecular tests using the Verigene Gram-Positive and Gram-Negative Blood Culture nucleic acid tests (BC-GP and BC-GN, resp.; Naosphere, Northbrook, IL, USA was evaluated for the identification of microorganisms detected from blood cultures. Ninety-nine blood cultures containing Gram-positive bacteria and 150 containing Gram-negative bacteria were analyzed using the BC-GP and BC-GN assays, respectively. Blood cultures were performed using the Bactec blood culture system (BD Diagnostic Systems, Franklin Lakes, NJ, USA and conventional identification and antibiotic-susceptibility tests were performed using a MicroScan system (Siemens, West Sacramento, CA, USA. When a single strain of bacteria was isolated from the blood culture, Verigene assays correctly identified 97.9% (94/96 of Gram-positive bacteria and 93.8% (137/146 of Gram-negative bacteria. Resistance genes mecA and vanA were correctly detected by the BC-GP assay, while the extended-spectrum β-lactamase CTX-M and the carbapenemase OXA resistance gene were detected from 30 cases cultures by the BC-GN assay. The BC-GP and BC-GN assays showed high agreement with conventional identification and susceptibility tests. These tests are useful for rapid identification of microorganisms and the detection of clinically important resistance genes from positive Bactec blood cultures.

  13. Evaluation of Verigene Blood Culture Test Systems for Rapid Identification of Positive Blood Cultures.

    Science.gov (United States)

    Kim, Jae-Seok; Kang, Go-Eun; Kim, Han-Sung; Kim, Hyun Soo; Song, Wonkeun; Lee, Kyu Man

    2016-01-01

    The performance of molecular tests using the Verigene Gram-Positive and Gram-Negative Blood Culture nucleic acid tests (BC-GP and BC-GN, resp.; Naosphere, Northbrook, IL, USA) was evaluated for the identification of microorganisms detected from blood cultures. Ninety-nine blood cultures containing Gram-positive bacteria and 150 containing Gram-negative bacteria were analyzed using the BC-GP and BC-GN assays, respectively. Blood cultures were performed using the Bactec blood culture system (BD Diagnostic Systems, Franklin Lakes, NJ, USA) and conventional identification and antibiotic-susceptibility tests were performed using a MicroScan system (Siemens, West Sacramento, CA, USA). When a single strain of bacteria was isolated from the blood culture, Verigene assays correctly identified 97.9% (94/96) of Gram-positive bacteria and 93.8% (137/146) of Gram-negative bacteria. Resistance genes mecA and vanA were correctly detected by the BC-GP assay, while the extended-spectrum β-lactamase CTX-M and the carbapenemase OXA resistance gene were detected from 30 cases cultures by the BC-GN assay. The BC-GP and BC-GN assays showed high agreement with conventional identification and susceptibility tests. These tests are useful for rapid identification of microorganisms and the detection of clinically important resistance genes from positive Bactec blood cultures.

  14. Rapid urine preparation prior to identification of uropathogens by MALDI-TOF MS.

    Science.gov (United States)

    Veron, L; Mailler, S; Girard, V; Muller, B H; L'Hostis, G; Ducruix, C; Lesenne, A; Richez, A; Rostaing, H; Lanet, V; Ghirardi, S; van Belkum, A; Mallard, F

    2015-09-01

    Matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-ToF MS) has been introduced in clinical routine microbiology laboratories. For the rapid diagnosis of urinary tract infections, culture-independent methods prior MALDI-mediated identification have been described. Here, we describe a comparison of three of these methods based on their performance of bacterial identification and their potential as a routine tool for microbiology labs : (i) differential centrifugation, (ii) urine filtration and (iii) a 5-h bacterial cultivation on solid culture media. For 19 urine samples, all methods were directly compared and correct bacterial species identification by MALDI was used as performance indicator. A higher percentage of correct MALDI identification was obtained after filtration (78.9 %) and the growth-based method (84.2 %) as compared to differential centrifugation (68.4 %). Additional testing of 76 mono-microbial specimens (bacteriuria > 10(5) CFU/mL) confirmed the good performance of short growth with a 90.8 % correct MALDI score, with a potentially better fit to the routine workflow of microbiology labs.

  15. Rapid experimental SAD phasing and hot-spot identification with halogenated fragments

    Directory of Open Access Journals (Sweden)

    Joseph D. Bauman

    2016-01-01

    Full Text Available Through X-ray crystallographic fragment screening, 4-bromopyrazole was discovered to be a `magic bullet' that is capable of binding at many of the ligand `hot spots' found in HIV-1 reverse transcriptase (RT. The binding locations can be in pockets that are `hidden' in the unliganded crystal form, allowing rapid identification of these sites for in silico screening. In addition to hot-spot identification, this ubiquitous yet specific binding provides an avenue for X-ray crystallographic phase determination, which can be a significant bottleneck in the determination of the structures of novel proteins. The anomalous signal from 4-bromopyrazole or 4-iodopyrazole was sufficient to determine the structures of three proteins (HIV-1 RT, influenza A endonuclease and proteinase K by single-wavelength anomalous dispersion (SAD from single crystals. Both compounds are inexpensive, readily available, safe and very soluble in DMSO or water, allowing efficient soaking into crystals.

  16. Rapid identification and differentiation of Saccharomyces cerevisiae, Saccharomyces bayanus and their hybrids by multiplex PCR.

    Science.gov (United States)

    Torriani, S; Zapparoli, G; Malacrinò, P; Suzzi, G; Dellaglio, F

    2004-01-01

    To develop a multiplex PCR assay for the specific identification and differentiation of Saccharomyces cerevisiae, S. bayanus and their hybrids. Two sets of primers with sequences complementary to the region YBR033w were used. A single amplicon of 1710 bp or 329 bp was obtained with species S. cerevisiae and S. bayanus, respectively, while the presence of both bands was observed in S. pastorianus because of its hybrid nature. Both amplification products were also obtained after amplification from DNA of several laboratory S. cerevisiae x S. bayanus hybrid strains. Multiplex PCR was optimized for the rapid and reliable identification of S. cerevisiae, S. bayanus and their hybrids. The procedure may be used for routine detection of the most common Saccharomyces sensu stricto yeasts involved in industrial fermentation processes, overcoming the problems of conventional techniques.

  17. Bacteriophage-based latex agglutination test for rapid identification of Staphylococcus aureus.

    Science.gov (United States)

    Idelevich, Evgeny A; Walther, Thomas; Molinaro, Sonja; Li, Xuehua; Xia, Guoqing; Wieser, Andreas; Peters, Georg; Peschel, Andreas; Becker, Karsten

    2014-09-01

    Rapid diagnosis is essential for the management of Staphylococcus aureus infections. A host recognition protein from S. aureus bacteriophage phiSLT was recombinantly produced and used to coat streptavidin latex beads to develop a latex agglutination test (LAT). The diagnostic accuracy of this bacteriophage-based test was compared with that of a conventional LAT, Pastorex Staph-Plus, by investigating a clinical collection of 86 S. aureus isolates and 128 coagulase-negative staphylococci (CoNS) from deep tissue infections. All of the clinical S. aureus isolates were correctly identified by the bacteriophage-based test. While most of the CoNS were correctly identified as non-S. aureus isolates, 7.9% of the CoNS caused agglutination. Thus, the sensitivity of the bacteriophage-based LAT for identification of S. aureus among clinical isolates was 100%, its specificity was 92.1%, its positive predictive value (PPV) was 89.6%, and its negative predictive value (NPV) was 100%. The sensitivity, specificity, PPV, and NPV of the Pastorex LAT for the identification of S. aureus were 100%, 99.2%, 98.9%, and 100%, respectively. Among the additionally tested 35 S. aureus and 91 non-S. aureus staphylococcal reference and type strains, 1 isolate was false negative by each system; 13 and 8 isolates were false positive by the bacteriophage-based and Pastorex LATs, respectively. The ability of the phiSLT protein to detect S. aureus was dependent on the presence of wall teichoic acid (WTA) and corresponded to the production of ribitol phosphate WTA, which is found in most S. aureus clones but only a small minority of CoNS. Bacteriophage-based LAT identification is a promising strategy for rapid pathogen identification. Finding more specific bacteriophage proteins would increase the specificity of this novel diagnostic approach. Copyright © 2014, American Society for Microbiology. All Rights Reserved.

  18. A rapid MALDI-TOF MS identification database at genospecies level for clinical and environmental Aeromonas strains.

    Directory of Open Access Journals (Sweden)

    Cinzia Benagli

    Full Text Available The genus Aeromonas has undergone a number of taxonomic and nomenclature revisions over the past 20 years, and new (subspecies and biogroups are continuously described. Standard identification methods such as biochemical characterization have deficiencies and do not allow clarification of the taxonomic position. This report describes the development of a matrix-assisted laser desorption/ionisation-time of flight mass spectrometry (MALDI-TOF MS identification database for a rapid identification of clinical and environmental Aeromonas isolates.

  19. Rapid identification of Stenotrophomonas maltophilia by peptide nucleic acid fluorescence in situ hybridization.

    Science.gov (United States)

    Hansen, N; Rasmussen, A K I; Fiandaca, M J; Kragh, K N; Bjarnsholt, T; Høiby, N; Stender, H; Guardabassi, L

    2014-05-01

    The objective of this study was to develop a novel peptide nucleic acid (PNA) probe for Stenotrophomonas maltophilia identification by fluorescence in situ hybridization (FISH). The probe was evaluated using 33 human and veterinary clinical S. maltophilia isolates and 45 reference strains representing common bacterial species in the respiratory tract. The probe displayed 100% sensitivity and 100% specificity on pure cultures and allowed detection in sputum from cystic fibrosis patients. The detection limit was 10(4) CFU/mL in spiked tracheal aspirate and bronchoalveolar lavage from healthy horses. Altogether the study shows that this species-specific PNA FISH probe facilitates rapid detection of S. maltophilia in biological specimens.

  20. Seed oil polyphenols: rapid and sensitive extraction method and high resolution-mass spectrometry identification.

    Science.gov (United States)

    Koubaa, Mohamed; Mhemdi, Houcine; Vorobiev, Eugène

    2015-05-01

    Phenolic content is a primary parameter for vegetables oil quality evaluation, and directly involved in the prevention of oxidation and oil preservation. Several methods have been reported in the literature for polyphenols extraction from seed oil but the approaches commonly used remain manually handled. In this work, we propose a rapid and sensitive method for seed oil polyphenols extraction and identification. For this purpose, polyphenols were extracted from Opuntia stricta Haw seed oil, using high frequency agitation, separated, and then identified using a liquid chromatography-high resolution mass spectrometry method. Our results showed good sensitivity and reproducibility of the developed methods. Copyright © 2015 Elsevier Inc. All rights reserved.

  1. Identification of the Species of Origin for Meat Products by Rapid Evaporative Ionization Mass Spectrometry.

    Science.gov (United States)

    Balog, Julia; Perenyi, Dora; Guallar-Hoyas, Cristina; Egri, Attila; Pringle, Steven D; Stead, Sara; Chevallier, Olivier P; Elliott, Chris T; Takats, Zoltan

    2016-06-15

    Increasingly abundant food fraud cases have brought food authenticity and safety into major focus. This study presents a fast and effective way to identify meat products using rapid evaporative ionization mass spectrometry (REIMS). The experimental setup was demonstrated to be able to record a mass spectrometric profile of meat specimens in a time frame of <5 s. A multivariate statistical algorithm was developed and successfully tested for the identification of animal tissue with different anatomical origin, breed, and species with 100% accuracy at species and 97% accuracy at breed level. Detection of the presence of meat originating from a different species (horse, cattle, and venison) has also been demonstrated with high accuracy using mixed patties with a 5% detection limit. REIMS technology was found to be a promising tool in food safety applications providing a reliable and simple method for the rapid characterization of food products.

  2. Rapid Detection and Identification of Yersinia pestis from Food Using Immunomagnetic Separation and Pyrosequencing

    Directory of Open Access Journals (Sweden)

    Kingsley K. Amoako

    2012-01-01

    Full Text Available Interest has recently been renewed in the possible use of Y. pestis, the causative agent of plague, as a biological weapon by terrorists. The vulnerability of food to intentional contamination coupled with reports of humans having acquired plague through eating infected animals that were not adequately cooked or handling of meat from infected animals makes the possible use of Y. pestis in a foodborne bioterrorism attack a reality. Rapid, efficient food sample preparation and detection systems that will help overcome the problem associated with the complexity of the different matrices and also remove any ambiguity in results will enable rapid informed decisions to be made regarding contamination of food with biothreat agents. We have developed a rapid detection assay that combines the use of immunomagnetic separation and pyrosequencing in generating results for the unambiguous identification of Y. pestis from milk (0.9 CFU/mL, bagged salad (1.6 CFU/g, and processed meat (10 CFU/g. The low detection limits demonstrated in this assay provide a novel tool for the rapid detection and confirmation of Y. pestis in food without the need for enrichment. The combined use of the iCropTheBug system and pyrosequencing for efficient capture and detection of Y. pestis is novel and has potential applications in food biodefence.

  3. Rapid identification of ubiquitination and SUMOylation target sites by microfluidic peptide array

    Directory of Open Access Journals (Sweden)

    Bing Zhu

    2016-03-01

    Full Text Available SUMOylation and ubiquitination are two essential post translational modifications (PTMs involved in the regulation of important biological processes in eukaryotic cells. Identification of ubiquitin (Ub and small ubiquitin-related modifier (SUMO-conjugated lysine residues in proteins is critical for understanding the role of ubiquitination and SUMOylation, but remains experimentally challenging. We have developed a powerful in vitro Ub/SUMO assay using a novel high density peptide array incorporated within a microfluidic device that allows rapid identification of ubiquitination and SUMOylation sites on target proteins. We performed the assay with a panel of human proteins and a microbial effector with known target sites for Ub or SUMO modifications, and determined that 80% of these proteins were modified by Ub or specific SUMO isoforms at the sites previously determined using conventional methods. Our results confirm the specificity for both SUMO isoform and individual target proteins at the peptide level. In summary, this microfluidic high density peptide array approach is a rapid screening assay to determine sites of Ub and SUMO modification of target substrates, which will provide new insights into the composition, selectivity and specificity of these PTM target sites.

  4. [Rapid identification of microorganisms by mass spectrometry in a blood culture system. Comparison of two procedures].

    Science.gov (United States)

    Cattani, María E; Posse, Tamara; Hermes, Ricardo L; Kaufman, Sara C

    2015-01-01

    Rapid identification of microorganisms is critical in hospitalized infected patients. Blood culture is currently the gold standard for detecting and identifying microorganisms causing bacteremia or sepsis. The introduction of mass spectrometry by matrix-assisted laser desorption/ionization time-of-flight (MALDI-TOF MS) in microbiology laboratories, especially in microorganisms growing in blood culture bottles, provides rapid identification. This study evaluates the performance of the Maldi Sepsityper Biotyper procedure (hereinafter, MS) compared to that of an in-home method (hereinafter, HF). Eight hundred and forty (840) positive blood culture bottles were processed using the HF procedure, 542 of which were also processed using MS. The organisms were identified in 670 (79.76%) and 391 (72.14%) bottles respectively (p = 0,0013). This study demonstrates the effectiveness of both procedures for identifying microorganisms directly from positive blood culture bottles. However, the HF procedure proved to be more effective than MS, especially in the presence of Gram positive organisms. Copyright © 2015 Asociación Argentina de Microbiología. Publicado por Elsevier España, S.L.U. All rights reserved.

  5. [Optinization of rapid propagation technique and induction and identification of autotetraploid of Polygonum multiflorum].

    Science.gov (United States)

    Huang, He-Ping; Gao, Shan-Lin; Wang, Jian; Huang, Lu-Qi; Huang, Peng

    2013-05-01

    To establish and optimize the rapid propagation system of Polygonum multiflorum, as well as explore method for induction and identification of autotetraploid. Propagation medium was optimized by orthogonal test. The buds were immersed in colchicine solution with different concentrations for different time to select induction conditions for autotetraploid of P. multiflorum. The most appropriate propagation medium was MS medium supplemented with 1.0 mg x L(-1) 6-BA, 0.3 mg x L(-1) NAA, and 0.4 mg x L(-1) PP333. That the buds were soaked in 0.2% colchicine solution for 30 h, or soaked in 0.3% colchicine solution for 18 h, was optimal condition to induce autopolyploid of P. multiflorum with induction rate as high as 16.7%. Rapid propagation of P. multiflorum could be achieved by tissue culture. Furthermore, colchicine was an effective inducer of polyploidy, and 25 tetraploid lines were obtained through chromosome identification. The experiment laid a foundation for the wild resource conservation, superior varieties breeding of P. multiflorum.

  6. Rapid identification of ST131 Escherichia coli by a novel multiplex real-time allelic discrimination assay.

    Science.gov (United States)

    François, Patrice; Bonetti, Eve-Julie; Fankhauser, Carolina; Baud, Damien; Cherkaoui, Abdessalam; Schrenzel, Jacques; Harbarth, Stephan

    2017-09-01

    Escherichia coli sequence type 131 is increasingly described in severe hospital infections. We developed a rapid real-time allelic discrimination assay for the rapid identification of E. coli ST131 isolates. This rapid assay represents an affordable alternative to sequence-based strategies before completing characterization of potentially highly virulent isolates of E. coli. Copyright © 2017 Elsevier B.V. All rights reserved.

  7. Prioritized Identification of Attractive and Romantic Partner Faces in Rapid Serial Visual Presentation.

    Science.gov (United States)

    Nakamura, Koyo; Arai, Shihoko; Kawabata, Hideaki

    2017-11-01

    People are sensitive to facial attractiveness because it is an important biological and social signal. As such, our perceptual and attentional system seems biased toward attractive faces. We tested whether attractive faces capture attention and enhance memory access in an involuntary manner using a dual-task rapid serial visual presentation (dtRSVP) paradigm, wherein multiple faces were successively presented for 120 ms. In Experiment 1, participants (N = 26) were required to identify two female faces embedded in a stream of animal faces as distractors. The results revealed that identification of the second female target (T2) was better when it was attractive compared to neutral or unattractive. In Experiment 2, we investigated whether perceived attractiveness affects T2 identification (N = 27). To this end, we performed another dtRSVP task involving participants in a romantic partnership with the opposite sex, wherein T2 was their romantic partner's face. The results demonstrated that a romantic partner's face was correctly identified more often than was the face of a friend or unknown person. Furthermore, the greater the intensity of passionate love participants felt for their partner (as measured by the Passionate Love Scale), the more often they correctly identified their partner's face. Our experiments indicate that attractive and romantic partners' faces facilitate the identification of the faces in an involuntary manner.

  8. Frequency regularities of acoustic modes and multi-colour mode identification in rapidly rotating stars

    Science.gov (United States)

    Reese, D. R.; Lignières, F.; Ballot, J.; Dupret, M.-A.; Barban, C.; van't Veer-Menneret, C.; MacGregor, K. B.

    2017-05-01

    Context. Mode identification has remained a major obstacle in the interpretation of pulsation spectra in rapidly rotating stars. This has motivated recent work on calculating realistic multi-colour mode visibilities in this type of star. Aims: We would like to test mode identification methods and seismic diagnostics in rapidly rotating stars, using oscillation spectra that are based on these new theoretical predictions. Methods: We investigate the auto-correlation function and Fourier transform of theoretically calculated frequency spectra, in which modes are selected according to their visibilities. Given that intrinsic mode amplitudes are determined by non-linear saturation and cannot currently be theoretically predicted, we experimented with various ad-hoc prescriptions for setting the mode amplitudes, including using random values. Furthermore, we analyse the ratios between mode amplitudes observed in different photometric bands to see up to what extent they can identify modes. Results: When non-random intrinsic mode amplitudes are used, our results show that it is possible to extract a mean value for the large frequency separation or half its value and, sometimes, twice the rotation rate, from the auto-correlation of the frequency spectra. Furthermore, the Fourier transforms are mostly sensitive to the large frequency separation or half its value. The combination of the two methods may therefore measure and distinguish the two types of separations. When the intrinsic mode amplitudes include random factors, which seems more representative of real stars, the results are far less favourable. It is only when the large separation or half its value coincides with twice the rotation rate, that it might be possible to detect the signature of a frequency regularity. We also find that amplitude ratios are a good way of grouping together modes with similar characteristics. By analysing the frequencies of these groups, it is possible to constrain mode identification, as

  9. Rapid Electrochemical Detection and Identification of Microbiological and Chemical Contaminants for Manned Spaceflight Project

    Science.gov (United States)

    Pierson, Duane; Botkin, Douglas; Gazda, Daniel

    2014-01-01

    Microbial control in the spacecraft environment is a daunting task, especially in the presence of human crew members. Currently, assessing the potential crew health risk associated with a microbial contamination event requires return of representative environmental samples that are analyzed in a ground-based laboratory. It is therefore not currently possible to quickly identify microbes during spaceflight. This project addresses the unmet need for spaceflight-compatible microbial identification technology. The electrochemical detection and identification platform is expected to provide a sensitive, specific, and rapid sample-to-answer capability for in-flight microbial monitoring that can distinguish between related microorganisms (pathogens and non-pathogens) as well as chemical contaminants. This will dramatically enhance our ability to monitor the spacecraft environment and the health risk to the crew. Further, the project is expected to eliminate the need for sample return while significantly reducing crew time required for detection of multiple targets. Initial work will focus on the optimization of bacterial detection and identification. The platform is designed to release nucleic acids (DNA and RNA) from microorganisms without the use of harmful chemicals. Bacterial DNA or RNA is captured by bacteria-specific probe molecules that are bound to a microelectrode, and that capture event can generate a small change in the electrical current (Lam, et al. 2012. Anal. Chem. 84(1): 21-5.). This current is measured, and a determination is made whether a given microbe is present in the sample analyzed. Chemical detection can be accomplished by directly applying a sample to the microelectrode and measuring the resulting current change. This rapid microbial and chemical detection device is designed to be a low-cost, low-power platform anticipated to be operated independently of an external power source, characteristics optimal for manned spaceflight and areas where power

  10. Rapid detection and identification of human hookworm infections through high resolution melting (HRM analysis.

    Directory of Open Access Journals (Sweden)

    Romano Ngui

    Full Text Available BACKGROUND: Hookworm infections are still endemic in low and middle income tropical countries with greater impact on the socioeconomic and public health of the bottom billion of the world's poorest people. In this study, a real-time polymerase chain reaction (PCR coupled with high resolution melting-curve (HRM analysis was evaluated for an accurate, rapid and sensitive tool for species identification focusing on the five human hookworm species. METHODS: Real-time PCR coupled with HRM analysis targeting the second internal transcribed spacer (ITS-2 of nuclear ribosomal DNA as the genetic marker was used to identify and distinguish hookworm species in human samples. Unique and distinct characteristics of HRM patterns were produced for each of the five hookworm species. The melting curves were characterized by peaks of 79.24±0.05°C and 83.00±0.04°C for Necator americanus, 79.12±0.10°C for Ancylostoma duodenale, 79.40±0.10°C for Ancylostoma ceylanicum, 79.63±0.05°C for Ancylostoma caninum and 79.70±0.14°C for Ancylostoma braziliense. An evaluation of the method's sensitivity and specificity revealed that this assay was able to detect as low as 0.01 ng/µl hookworm DNA and amplification was only recorded for hookworm positive samples. CONCLUSION: The HRM assay developed in this study is a rapid and straightforward method for the diagnosis, identification and discrimination of five human hookworms. This assay is simple compared to other probe-based genotyping methods as it does not require multiplexing, DNA sequencing or post-PCR processing. Therefore, this method offers a new alternative for rapid detection of human hookworm species.

  11. Rapid detection and identification of human hookworm infections through high resolution melting (HRM) analysis.

    Science.gov (United States)

    Ngui, Romano; Lim, Yvonne A L; Chua, Kek Heng

    2012-01-01

    Hookworm infections are still endemic in low and middle income tropical countries with greater impact on the socioeconomic and public health of the bottom billion of the world's poorest people. In this study, a real-time polymerase chain reaction (PCR) coupled with high resolution melting-curve (HRM) analysis was evaluated for an accurate, rapid and sensitive tool for species identification focusing on the five human hookworm species. Real-time PCR coupled with HRM analysis targeting the second internal transcribed spacer (ITS-2) of nuclear ribosomal DNA as the genetic marker was used to identify and distinguish hookworm species in human samples. Unique and distinct characteristics of HRM patterns were produced for each of the five hookworm species. The melting curves were characterized by peaks of 79.24±0.05°C and 83.00±0.04°C for Necator americanus, 79.12±0.10°C for Ancylostoma duodenale, 79.40±0.10°C for Ancylostoma ceylanicum, 79.63±0.05°C for Ancylostoma caninum and 79.70±0.14°C for Ancylostoma braziliense. An evaluation of the method's sensitivity and specificity revealed that this assay was able to detect as low as 0.01 ng/µl hookworm DNA and amplification was only recorded for hookworm positive samples. The HRM assay developed in this study is a rapid and straightforward method for the diagnosis, identification and discrimination of five human hookworms. This assay is simple compared to other probe-based genotyping methods as it does not require multiplexing, DNA sequencing or post-PCR processing. Therefore, this method offers a new alternative for rapid detection of human hookworm species.

  12. Rapid Identification of Microorganisms by FilmArray Blood Culture Identification Panel Improves Clinical Management in Children.

    Science.gov (United States)

    Ray, Stephen T J; Drew, Richard J; Hardiman, Fiona; Pizer, Barry; Riordan, Andrew

    2016-05-01

    Blood cultures are a common investigation for children admitted to hospital. In routine practice, it takes at least 24 hours to identify an organism as a contaminant or clinically significant. FilmArray Blood Culture Identification Panel (FA-BCIP) is a multiplex polymerase chain reaction that can detect 24 pathogens within 1 hour. We assessed whether results from FA-BCIP lead to changes in clinical management in a tertiary referral paediatric hospital. We prospectively studied children having blood cultures taken at our tertiary children's hospital. Blood cultures were monitored and organisms identified using standard methods. FA-BCIP was performed when growth was initially detected in first positive blood cultures per episode, between January 1 and June 30, 2014. Assessment of whether the FA-BCIP result altered clinical management was made, specifically focused on antimicrobial stewardship and length of stay. FA-BCIP was done on 117 positive blood cultures; 74 (63%) grew clinically significant organisms, 43 (37%) grew contaminants. FA-BCIP results were judged to alter clinical management in 63 of the 117 episodes (54%). Antimicrobials were started/altered in 23 (19%) episodes and de-escalated/withheld/stopped in 29 (25%) episodes. Ten children were discharged from hospital earlier, which saved a cumulative total of 14 bed days. Rapid identification of microorganisms in pediatric blood cultures by FA-BCIP, led to changes in clinical management for half of the episodes. This improved antimicrobial stewardship and allowed early discharge from hospital for 10% of children. Future studies should focus on how best to use this technology in a cost-effective manner.

  13. Rapid Identification of Emerging Human-Pathogenic Sporothrix Species with Rolling Circle Amplification

    Science.gov (United States)

    Rodrigues, Anderson M.; Najafzadeh, Mohammad J.; de Hoog, G. Sybren; de Camargo, Zoilo P.

    2015-01-01

    Sporothrix infections are emerging as an important human and animal threat among otherwise healthy patients, especially in Brazil and China. Correct identification of sporotrichosis agents is beneficial for epidemiological surveillance, enabling implementation of adequate public-health policies and guiding antifungal therapy. In areas of limited resources where sporotrichosis is endemic, high-throughput detection methods that are specific and sensitive are preferred over phenotypic methods that usually result in misidentification of closely related Sporothrix species. We sought to establish rolling circle amplification (RCA) as a low-cost screening tool for species-specific identification of human-pathogenic Sporothrix. We developed six species-specific padlock probes targeting polymorphisms in the gene encoding calmodulin. BLAST-searches revealed candidate probes that were conserved intraspecifically; no significant homology with sequences from humans, mice, plants or microorganisms outside members of Sporothrix were found. The accuracy of our RCA-based assay was demonstrated through the specificity of probe-template binding to 25 S. brasiliensis, 58 S. schenckii, 5 S. globosa, 1 S. luriei, 4 S. mexicana, and 3 S. pallida samples. No cross reactivity between closely related species was evident in vitro, and padlock probes yielded 100% specificity and sensitivity down to 3 × 106 copies of the target sequence. RCA-based speciation matched identifications via phylogenetic analysis of the gene encoding calmodulin and the rDNA operon (kappa 1.0; 95% confidence interval 1.0-1.0), supporting its use as a reliable alternative to DNA sequencing. This method is a powerful tool for rapid identification and specific detection of medically relevant Sporothrix, and due to its robustness has potential for ecological studies. PMID:26696992

  14. A multiplex PCR for rapid identification of Brassica species in the triangle of U.

    Science.gov (United States)

    Koh, Joshua C O; Barbulescu, Denise M; Norton, Sally; Redden, Bob; Salisbury, Phil A; Kaur, Sukhjiwan; Cogan, Noel; Slater, Anthony T

    2017-01-01

    Within the Brassicaceae, six species from the genus Brassica are widely cultivated throughout the world as oilseed, condiment, fodder or vegetable crops. The genetic relationships among the six Brassica species are described by U's triangle model. Extensive shared traits and diverse morphotypes among Brassica species make identification and classification based on phenotypic data alone challenging and unreliable, especially when dealing with large germplasm collections. Consequently, a major issue for genebank collections is ensuring the correct identification of species. Molecular genotyping based on simple sequence repeat (SSR) marker sequencing or the Illumina Infinium Brassica napus 60K single nucleotide polymorphism (SNP) array has been used to identify species and assess genetic diversity of Brassica collections. However, these methods are technically challenging, expensive and time-consuming, making them unsuitable for routine or rapid screening of Brassica accessions for germplasm management. A cheaper, faster and simpler method for Brassica species identification is described here. A multiplex polymerase chain reaction (MPCR) consisting of new and existing primers specific to the Brassica A, B and C genomes was able to reliably distinguish all six Brassica species in the triangle of U with 16 control samples of known species identity. Further validation against 120 Brassica accessions previously genotyped showed that the MPCR is highly accurate and comparable to more advanced techniques such as SSR marker sequencing or the Illumina Infinium B. napus 60K SNP array. In addition, the MPCR was sensitive enough to detect seed contaminations in pooled seed samples of Brassica accessions. A cheap and fast multiplex PCR assay for identification of Brassica species in the triangle of U was developed and validated in this study. The MPCR assay can be readily implemented in any basic molecular laboratory and should prove useful for the management of Brassica

  15. Rapid detection and identification of viroids in the genus Coleviroid using a universal probe.

    Science.gov (United States)

    Jiang, Dongmei; Hou, Wanying; Sano, Teruo; Kang, Ni; Qin, Lü; Wu, Zujian; Li, Shifang; Xie, Lianhui

    2013-02-01

    A simple, low-cost hybridization assay using a universal DIG-labeled riboprobe for the rapid detection and identification of coleus viroids is presented. An octamer of 32-nucleotide sequence derived from the central conserved region (CCR) of viroids in the genus Coleviroid was used to develop a universal cRNA probe (8-central-conserved-region probe, 8CCR probe) for coleus viroids. Dot-blot hybridization assays demonstrated that the sensitivity of this probe was similar to specific probes for each CbVd, and Northern hybridization results revealed that at least four coleus viroids could be distinguished readily and simultaneously using the 8CCR probe. Batch detection assay showed that hybridization using the 8CCR probe can identify coleus viroids rapidly and effectively. This rapid and low-cost molecular hybridization technique is an effective way to survey the occurrence of coleus viroids, and has reference for the detection of other viroids and possibly viruses. Copyright © 2012 Elsevier B.V. All rights reserved.

  16. Rapid Detection and Identification of a Pathogen's DNA Using Phi29 DNA Polymerase

    Energy Technology Data Exchange (ETDEWEB)

    Xu, Y.; Dunn, J.; Gao, S.; Bruno, J. F.; Luft, B. J.

    2008-10-31

    Zoonotic pathogens including those transmitted by insect vectors are some of the most deadly of all infectious diseases known to mankind. A number of these agents have been further weaponized and are widely recognized as being potentially significant biothreat agents. We describe a novel method based on multiply-primed rolling circle in vitro amplification for profiling genomic DNAs to permit rapid, cultivation-free differential detection and identification of circular plasmids in infectious agents. Using Phi29 DNA polymerase and a two-step priming reaction we could reproducibly detect and characterize by DNA sequencing circular DNA from Borrelia burgdorferi B31 in DNA samples containing as little as 25 pg of Borrelia DNA amongst a vast excess of human DNA. This simple technology can ultimately be adapted as a sensitive method to detect specific DNA from both known and unknown pathogens in a wide variety of complex environments.

  17. Rapid Identification of Methicillin-Resistant Staphylococcus aureus (MRSA) by the Vitek MS Saramis system.

    Science.gov (United States)

    Shan, Weiguang; Li, Jiaping; Fang, Ying; Wang, Xuan; Gu, Danxia; Zhang, Rong

    2016-01-01

    A rapid, sensitive, and accurate Vitek MS assay was developed to distinguish clinical isolates of methicillin-resistant Staphylococcus aureus (MRSA) from clinical isolates of methicillin-sensitive Staphylococcus aureus (MSSA) by developing an in-house knowledgebase of SuperSpectra. Three unique peaks, including peaks at 2305.6 and 3007.3 Da specific to MRSA, and 6816.7 Da specific to MSSA, were selected for differentiating MRSA and MSSA. This assay accurately identified 84 and 91% of clinical MRSA and MSSA strains out of the total 142 clinically acquired S. aureus strains that were tested. This method will greatly improve the efficiency of single clinical sample identification of MRSA, thereby facilitating a reduction in the transmission of MRSA in clinical settings.

  18. Rapid identification of three varieties of Chrysanthemum with near infrared spectroscopy

    Directory of Open Access Journals (Sweden)

    Cun-wu Chen

    Full Text Available A total of 139 batches of Chrysanthemum samples were randomly divided into calibration set (92 batches and prediction set (47 batches. The near infrared diffuses reflectance spectra of Chrysanthemum varieties were preprocessed by a first order derivative (D1 and autoscaling, and a modelwas built using partial least squares analysis. In this study, three Chrysanthemum varieties were identified, the accuracy rates in calibration sets of Dabaiju, Huju, and Xiaobaiju are 97.60, 96.65, and 94.70%, respectively; And 95.16, 86.11, and 93.46% accuracy rate in prediction sets was obtained. The research results demonstrate that the qualitative analysis can be conducted by machine learning combined with Near-Infrared Spectroscopy, which provides a new method for rapid and non-invasive identification of Chrysanthemum varieties.

  19. Multiplex Solid-Phase PCR for Rapid Detection and Identification of Salmonella spp. at Sub-species

    DEFF Research Database (Denmark)

    Cao, Cuong; Høgberg, Jonas; Wolff, Anders

    -PCR gel electrophoresis. The method will be useful for development of point-of-care devices for rapid detection and identification of Salmonella spp. A solid-phase PCR for rapid detection and identification of S. enteritidis, S. typhimurium and S. dublin is developed. The method offers advantages......This study presents a solid-phase PCR (SP-PCR) for rapid detection, identification, and sub-typing of various Salmonella species, the major food-borne cause of salmonellosis. The target DNA is firstly amplified with PCR primers (one primer is labeled with fluorophores) in the liquid phase...... by the liquid phase primer thus generating new templates for the SP-PCR. After the reaction, PCR products labeled with fluorophores remain attached to the substrate and can be visualized directly by fluorescence readout devices. Using this method, S. enteritidis, S. typhimurium and S. dublin can be detected...

  20. A novel biosensor for rapid identification of high temperature resistant species

    Science.gov (United States)

    Zhang, Lingrui; Xing, Da; Zhou, Xiaoming

    2007-11-01

    In this paper, a novel biosensor technique for identification of high temperature resistant species based on quantitative measurement of delayed fluorescence (DF) is described. The biosensor, which uses light-emitting diode lattice as excitation light source, is portable and can detect DF emission from plants in vivo. Compared with its primary version in our previous report, the biosensor presented here can better control environmental factors. Moreover, the improved biosensor can automatically complete the measurements of light response curves of DF intensity in a programmed mode. The testing of the improved biosensor has been made in two maize species (Zea May L.) after high temperature treatment. Contrast evaluations of the effects of heat stress on seedlings photosynthesis were made from measurements of net photosynthesis rate (Pn) based on consumption of CO II. Current testing has demonstrated that the DF intensity well correlates with Pn in each plant species after heat stress. We thus conclude that the DF technique is a breakthrough to traditional strategy of identifying the differences in heat tolerance based on gas exchange, and can provide a reliable approach for rapid and non-invasive determination of the effects of heat stress on photosynthesis and identification of high temperature resistant species.

  1. Rapid and high-throughput pan-Orthopoxvirus detection and identification using PCR and mass spectrometry.

    Directory of Open Access Journals (Sweden)

    Mark W Eshoo

    2009-07-01

    Full Text Available The genus Orthopoxvirus contains several species of related viruses, including the causative agent of smallpox (Variola virus. In addition to smallpox, several other members of the genus are capable of causing human infection, including monkeypox, cowpox, and other zoonotic rodent-borne poxviruses. Therefore, a single assay that can accurately identify all orthopoxviruses could provide a valuable tool for rapid broad orthopovirus identification. We have developed a pan-Orthopoxvirus assay for identification of all members of the genus based on four PCR reactions targeting Orthopoxvirus DNA and RNA helicase and polymerase genes. The amplicons are detected using electrospray ionization-mass spectrometry (PCR/ESI-MS on the Ibis T5000 system. We demonstrate that the assay can detect and identify a diverse collection of orthopoxviruses, provide sub-species information and characterize viruses from the blood of rabbitpox infected rabbits. The assay is sensitive at the stochastic limit of PCR and detected virus in blood containing approximately six plaque-forming units per milliliter from a rabbitpox virus-infected rabbit.

  2. Rapid and Direct VHH and Target Identification by Staphylococcal Surface Display Libraries

    Directory of Open Access Journals (Sweden)

    Marco Cavallari

    2017-07-01

    Full Text Available Unbiased and simultaneous identification of a specific antibody and its target antigen has been difficult without prior knowledge of at least one interaction partner. Immunization with complex mixtures of antigens such as whole organisms and tissue extracts including tumoral ones evokes a highly diverse immune response. During such a response, antibodies are generated against a variety of epitopes in the mixture. Here, we propose a surface display design that is suited to simultaneously identify camelid single domain antibodies and their targets. Immune libraries of single-domain antigen recognition fragments from camelid heavy chain-only antibodies (VHH were attached to the peptidoglycan of Gram-positive Staphylococcus aureus employing its endogenous housekeeping sortase enzyme. The sortase transpeptidation reaction covalently attached the VHH to the bacterial peptidoglycan. The reversible nature of the reaction allowed the recovery of the VHH from the bacterial surface and the use of the VHH in downstream applications. These staphylococcal surface display libraries were used to rapidly identify VHH as well as their targets by immunoprecipitation (IP. Our novel bacterial surface display platform was stable under harsh screening conditions, allowed fast target identification, and readily permitted the recovery of the displayed VHH for downstream analysis.

  3. Rapid experimental SAD phasing and hot-spot identification with halogenated fragments

    Energy Technology Data Exchange (ETDEWEB)

    Bauman, Joseph D.; Harrison, Jerry Joe E. K.; Arnold, Eddy

    2016-01-01

    Through X-ray crystallographic fragment screening, 4-bromopyrazole was discovered to be a `magic bullet' that is capable of binding at many of the ligand `hot spots' found in HIV-1 reverse transcriptase (RT). The binding locations can be in pockets that are `hidden' in the unliganded crystal form, allowing rapid identification of these sites forin silicoscreening. In addition to hot-spot identification, this ubiquitous yet specific binding provides an avenue for X-ray crystallographic phase determination, which can be a significant bottleneck in the determination of the structures of novel proteins. The anomalous signal from 4-bromopyrazole or 4-iodopyrazole was sufficient to determine the structures of three proteins (HIV-1 RT, influenza A endonuclease and proteinase K) by single-wavelength anomalous dispersion (SAD) from single crystals. Both compounds are inexpensive, readily available, safe and very soluble in DMSO or water, allowing efficient soaking into crystals.

  4. Rapid identification of rice blast resistance gene by specific length amplified fragment sequencing

    Directory of Open Access Journals (Sweden)

    Shen Chen

    2016-05-01

    Full Text Available Excavation of resistance genes is one of the most effective and environment-friendly measures to control the devastating rice disease caused by Magnaporthe oryzae. Many resistance genes have been mapped and characterized in the last century. Nevertheless, only a few of the total resistance genes could be really applied in the rice breeding program. Huazhan (HZ is a new native rice restorer line developed in China and widely used in hybrid rice in recent years. HZ and its crossed combinations usually show a broad spectrum of resistance against rice blast in different rice ecosystems in China. Dissection of the genetic background of HZ is very useful for its further application. In this study, a combined method based on bulked segregation analysis (BSA and specific length amplified fragment sequencing (SLAF-seq was used to identify blast resistance gene(s in HZ. A total of 56,187 SLAFs labels were captured and 9051 polymorphic SLAFs markers were analysed and procured in this study. One trait associated with candidate resistance genes region on chromosome 12 overlapping 10.2–17.6 Mb has been identified, in which 10 NBS-LRR (nucleotide-binding site-leucine-rich repeat coding genes were used as resistance gene candidates. Our result indicated that SLAF-seq with BSA is a rapid and effective method for initial identification of blast resistance genes. The identification of resistance gene in HZ will improve its molecular breeding and resistance variety application.

  5. Rapid and Direct VHH and Target Identification by Staphylococcal Surface Display Libraries.

    Science.gov (United States)

    Cavallari, Marco

    2017-07-12

    Unbiased and simultaneous identification of a specific antibody and its target antigen has been difficult without prior knowledge of at least one interaction partner. Immunization with complex mixtures of antigens such as whole organisms and tissue extracts including tumoral ones evokes a highly diverse immune response. During such a response, antibodies are generated against a variety of epitopes in the mixture. Here, we propose a surface display design that is suited to simultaneously identify camelid single domain antibodies and their targets. Immune libraries of single-domain antigen recognition fragments from camelid heavy chain-only antibodies (VHH) were attached to the peptidoglycan of Gram-positive Staphylococcus aureus employing its endogenous housekeeping sortase enzyme. The sortase transpeptidation reaction covalently attached the VHH to the bacterial peptidoglycan. The reversible nature of the reaction allowed the recovery of the VHH from the bacterial surface and the use of the VHH in downstream applications. These staphylococcal surface display libraries were used to rapidly identify VHH as well as their targets by immunoprecipitation (IP). Our novel bacterial surface display platform was stable under harsh screening conditions, allowed fast target identification, and readily permitted the recovery of the displayed VHH for downstream analysis.

  6. Rapid annotation of anonymous sequences from genome projects using semantic similarities and a weighting scheme in gene ontology.

    Directory of Open Access Journals (Sweden)

    Paolo Fontana

    Full Text Available BACKGROUND: Large-scale sequencing projects have now become routine lab practice and this has led to the development of a new generation of tools involving function prediction methods, bringing the latter back to the fore. The advent of Gene Ontology, with its structured vocabulary and paradigm, has provided computational biologists with an appropriate means for this task. METHODOLOGY: We present here a novel method called ARGOT (Annotation Retrieval of Gene Ontology Terms that is able to process quickly thousands of sequences for functional inference. The tool exploits for the first time an integrated approach which combines clustering of GO terms, based on their semantic similarities, with a weighting scheme which assesses retrieved hits sharing a certain number of biological features with the sequence to be annotated. These hits may be obtained by different methods and in this work we have based ARGOT processing on BLAST results. CONCLUSIONS: The extensive benchmark involved 10,000 protein sequences, the complete S. cerevisiae genome and a small subset of proteins for purposes of comparison with other available tools. The algorithm was proven to outperform existing methods and to be suitable for function prediction of single proteins due to its high degree of sensitivity, specificity and coverage.

  7. Improved protocol for rapid identification of certain spa types using high resolution melting curve analysis.

    Science.gov (United States)

    Mayerhofer, Benjamin; Stöger, Anna; Pietzka, Ariane T; Fernandez, Haizpea Lasa; Prewein, Bernhard; Sorschag, Sieglinde; Kunert, Renate; Allerberger, Franz; Ruppitsch, Werner

    2015-01-01

    Methicillin-resistant Staphylococcus aureus is one of the most significant pathogens associated with health care. For efficient surveillance, control and outbreak investigation, S. aureus typing is essential. A high resolution melting curve analysis was developed and evaluated for rapid identification of the most frequent spa types found in an Austrian hospital consortium covering 2,435 beds. Among 557 methicillin-resistant Staphylococcus aureus isolates 38 different spa types were identified by sequence analysis of the hypervariable region X of the protein A gene (spa). Identification of spa types through their characteristic high resolution melting curve profiles was considerably improved by double spiking with genomic DNA from spa type t030 and spa type t003 and allowed unambiguous and fast identification of the ten most frequent spa types t001 (58%), t003 (12%), t190 (9%), t041 (5%), t022 (2%), t032 (2%), t008 (2%), t002 (1%), t5712 (1%) and t2203 (1%), representing 93% of all isolates within this hospital consortium. The performance of the assay was evaluated by testing samples with unknown spa types from the daily routine and by testing three different high resolution melting curve analysis real-time PCR instruments. The ten most frequent spa types were identified from all samples and on all instruments with 100% specificity and 100% sensitivity. Compared to classical spa typing by sequence analysis, this gene scanning assay is faster, cheaper and can be performed in a single closed tube assay format. Therefore it is an optimal screening tool to detect the most frequent endemic spa types and to exclude non-endemic spa types within a hospital.

  8. Rapid detection and identification of four major Schistosoma species by high-resolution melt (HRM) analysis.

    Science.gov (United States)

    Li, Juan; Zhao, Guang-Hui; Lin, RuiQing; Blair, David; Sugiyama, Hiromu; Zhu, Xing-Quan

    2015-11-01

    Schistosomiasis, caused by blood flukes belonging to several species of the genus Schistosoma, is a serious and widespread parasitic disease. Accurate and rapid differentiation of these etiological agents of animal and human schistosomiasis to species level can be difficult. We report a real-time PCR assay coupled with a high-resolution melt (HRM) assay targeting a portion of the nuclear 18S rDNA to detect, identify, and distinguish between four major blood fluke species (Schistosoma japonicum, Schistosoma mansoni, Schistosoma haematobium, and Schistosoma mekongi). Using this system, the Schistosoma spp. was accurately identified and could also be distinguished from all other trematode species with which they were compared. As little as 10(-5) ng genomic DNA from a Schistosoma sp. could be detected. This process is inexpensive, easy, and can be completed within 3 h. Examination of 21 representative Schistosoma samples from 15 geographical localities in seven endemic countries validated the value of the HRM detection assay and proved its reliability. The melting curves were characterized by peaks of 83.65 °C for S. japonicum and S. mekongi, 85.65 °C for S. mansoni, and 85.85 °C for S. haematobium. The present study developed a real-time PCR coupled with HRM analysis assay for detection and differential identification of S. mansoni, S. haematobium, S. japonicum, and S. mekongi. This method is rapid, sensitive, and inexpensive. It has important implications for epidemiological studies of Schistosoma.

  9. FT-IR microspectroscopy in rapid identification of bacteria in pure and mixed culture

    Science.gov (United States)

    Fontoura, Inglid; Belo, Ricardo; Sakane, Kumiko; Cardoso, Maria Angélica Gargione; Khouri, Sônia; Uehara, Mituo; Raniero, Leandro; Martin, Airton A.

    2010-02-01

    In recent years FT-IR microspectroscopy has been developed for microbiology analysis and applied successfully in pure cultures of microorganisms to rapidly identify strains of bacteria, yeasts and fungi. The investigation and characterization of microorganism mixed cultures is also of growing importance, especially in hospitals where it is common to poly-microbial infections. In this work, the rapid identification of bacteria in pure and mixed cultures was studied. The bacteria were obtained from the Institute Oswaldo Cruz culture collection at Brazil. Escherichia coli ATCC 10799 and Staphylococcus aureus ATCC 14456 were analyzed, 3 inoculations were examined in triplicate: Escherichia coli, Staphylococcus aureus and a mixed culture of them. The inoculations were prepared according to McFarland 0.5, incubated at 37 ° C for 6 hours, diluted in saline, placed in the CaF2 window and store for one hour at 50°C to obtain thin film. The measurement was performed by Spectrum Spotlight 400 (Perkin-Elmer) equipment in the range of 4000-900 cm-1, with 32 scans using a transmittance technique with point and image modes. The data were processed (baseline, normalization, calculation of first derivate followed by smoothing with 9 point using a Savitzky-Golay algorithm) and a cluster analysis were done by Ward's algorithm and an excellent discrimination between pure and mixed culture was obtained. Our preliminary results indicate that the FT-IR microspectroscopy associated with cluster analysis can be used to discriminate between pure and mixed culture.

  10. Rapid identification of antifungal compounds against Exserohilum rostratum using high throughput drug repurposing screens.

    Science.gov (United States)

    Sun, Wei; Park, Yoon-Dong; Sugui, Janyce A; Fothergill, Annette; Southall, Noel; Shinn, Paul; McKew, John C; Kwon-Chung, Kyung J; Zheng, Wei; Williamson, Peter R

    2013-01-01

    A recent large outbreak of fungal infections by Exserohilum rostratum from contaminated compounding solutions has highlighted the need to rapidly screen available pharmaceuticals that could be useful in therapy. The present study utilized two newly-developed high throughput assays to screen approved drugs and pharmaceutically active compounds for identification of potential antifungal agents. Several known drugs were found that have potent effects against E. rostratum including the triazole antifungal posaconazole. Posaconazole is likely to be effective against infections involving septic joints and may provide an alternative for refractory central nervous system infections. The anti-E. rostratum activities of several other drugs including bithionol (an anti-parasitic drug), tacrolimus (an immunosuppressive agent) and floxuridine (an antimetabolite) were also identified from the drug repurposing screens. In addition, activities of other potential antifungal agents against E. rostratum were excluded, which may avoid unnecessary therapeutic trials and reveals the limited therapeutic alternatives for this outbreak. In summary, this study has demonstrated that drug repurposing screens can be quickly conducted within a useful time-frame. This would allow clinical implementation of identified alternative therapeutics and should be considered as part of the initial public health response to new outbreaks or rapidly-emerging microbial pathogens.

  11. Rapid identification of antifungal compounds against Exserohilum rostratum using high throughput drug repurposing screens.

    Directory of Open Access Journals (Sweden)

    Wei Sun

    Full Text Available A recent large outbreak of fungal infections by Exserohilum rostratum from contaminated compounding solutions has highlighted the need to rapidly screen available pharmaceuticals that could be useful in therapy. The present study utilized two newly-developed high throughput assays to screen approved drugs and pharmaceutically active compounds for identification of potential antifungal agents. Several known drugs were found that have potent effects against E. rostratum including the triazole antifungal posaconazole. Posaconazole is likely to be effective against infections involving septic joints and may provide an alternative for refractory central nervous system infections. The anti-E. rostratum activities of several other drugs including bithionol (an anti-parasitic drug, tacrolimus (an immunosuppressive agent and floxuridine (an antimetabolite were also identified from the drug repurposing screens. In addition, activities of other potential antifungal agents against E. rostratum were excluded, which may avoid unnecessary therapeutic trials and reveals the limited therapeutic alternatives for this outbreak. In summary, this study has demonstrated that drug repurposing screens can be quickly conducted within a useful time-frame. This would allow clinical implementation of identified alternative therapeutics and should be considered as part of the initial public health response to new outbreaks or rapidly-emerging microbial pathogens.

  12. A Simple and Rapid Identification Method for Mycobacterium bovis BCG with Loop-Mediated Isothermal Amplification.

    Directory of Open Access Journals (Sweden)

    Yuji Kouzaki

    Full Text Available Bacillus Calmette-Guérin (BCG is widely used as a live attenuated vaccine against Mycobacterium tuberculosis and is an agent for standard prophylaxis against the recurrence of bladder cancer. Unfortunately, it can cause severe infectious diseases, especially in immunocompromised patients, and the ability to immediately distinguish BCG from other M. tuberculosis complexes is therefore important. In this study, we developed a simple and easy-to-perform identification procedure using loop-mediated amplification (LAMP to detect deletions within the region of difference, which is deleted specifically in all M. bovis BCG strains. Reactions were performed at 64 °C for 30 min and successful targeted gene amplifications were detected by real-time turbidity using a turbidimeter and visual inspection of color change. The assay had an equivalent detection limit of 1.0 pg of genomic DNA using a turbidimeter whereas it was 10 pg with visual inspection, and it showed specificity against 49 strains of 44 pathogens, including M. tuberculosis complex. The expected LAMP products were confirmed through identical melting curves in real-time LAMP procedures. We employed the Procedure for Ultra Rapid Extraction (PURE kit to isolate mycobacterial DNA and found that the highest sensitivity limit with a minimum total cell count of mycobacterium (including DNA purification with PURE was up to 1 × 10(3 cells/reaction, based on color changes under natural light with FDA reagents. The detection limit of this procedure when applied to artificial serum, urine, cerebrospinal fluid, and bronchoalveolar lavage fluid samples was also about 1 × 10(3 cells/reaction. Therefore, this substitute method using conventional culture or clinical specimens followed by LAMP combined with PURE could be a powerful tool to enable the rapid identification of M. bovis BCG as point-of-care testing. It is suitable for practical use not only in resource-limited situations, but also in any

  13. [Automated RNA amplification for the rapid identification of Mycobacterium tuberculosis complex in respiratory specimens].

    Science.gov (United States)

    Drouillon, V; Houriez, F; Buze, M; Lagrange, P; Herrmann, J-L

    2006-01-01

    Rapid and sensitive detection of Mycobacterium tuberculosis complex (MTB) directly on clinical respiratory specimens is essential for a correct management of patients suspected of tuberculosis. For this purpose PCR-based kits are available to detect MTB in respiratory specimen but most of them need at least 4 hours to be completed. New methods, based on TRC method (TRC: Transcription Reverse transcription Concerted--TRCRapid M. Tuberculosis--Tosoh Bioscience, Tokyo, Japon) and dedicated monitor have been developed. A new kit (TRC Rapid M. tuberculosis and Real-time monitor TRCRapid-160, Tosoh Corporation, Japan) enabling one step amplification and real-time detection of MTB 16S rRNA by a combination of intercalative dye oxazole yellow-linked DNA probe and isothermal RNA amplification directly on respiratory specimens has been tested in our laboratory. 319 respiratory specimens were tested in this preliminary study and results were compared to smear and culture. Fourteen had a positive culture for MTB. Among theses samples, smear was positive in 11 cases (78.6%) and TRC process was positive in 8 cases (57.1%). Overall sensitivity of TRC compared to smear positive samples is 73%. Theses first results demonstrated that a rapid identification of MTB was possible (less than 2 processing hours for 14 specimens and about 1 hour for 1 specimen) in most cases of smear positive samples using ready to use reagents for real time detection of MTB rRNA in clinical samples. New pretreatment and extraction reagents kits to increase the stability of the sputum RNA and the extraction efficiency are now tested in our laboratory.

  14. Rapid identification of bacteria and candida using pna-fish from blood and peritoneal fluid cultures: a retrospective clinical study

    Directory of Open Access Journals (Sweden)

    Harris Dana M

    2013-01-01

    Full Text Available Abstract Background Peptide nucleic acid fluorescent in situ hybridization (PNA-FISH is a rapid and established method for identification of Candida sp., Gram positive, and Gram negative bacteria from positive blood cultures. This study reports clinical experience in the evaluation of 103 positive blood cultures and 17 positive peritoneal fluid cultures from 120 patients using PNA-FISH. Our study provides evidence as to potential pharmaceutical cost savings based on rapid pathogen identification, in addition to the novel application of PNA-FISH to peritoneal fluid specimens. Methods Identification accuracy and elapsed time to identification of Gram positives, Gram negatives, and Candida sp., isolated from blood and peritoneal fluid cultures were assessed using PNA-FISH (AdvanDx, as compared to standard culture methods. Patient charts were reviewed to extrapolate potential pharmaceutical cost savings due to adjustment of antimicrobial or antifungal therapy, based on identification by PNA-FISH. Results In blood cultures, time to identification by standard culture methods for bacteria and Candida sp., averaged 83.6 hours (95% CI 56.7 to 110.5. Identification by PNA-FISH averaged 11.2 hours (95% CI 4.8 to 17.6. Overall PNA-FISH identification accuracy was 98.8% (83/84, 95% CI 93.5% to 99.9% as compared to culture. In peritoneal fluid, identification of bacteria by culture averaged 87.4 hours (95% CI −92.4 to 267.1. Identification by PNA-FISH averaged 16.4 hours (95% CI −57.3 to 90.0. Overall PNA-FISH identification accuracy was 100% (13/13, 95% CI 75.3% to 100%. For Candida sp., pharmaceutical cost savings based on PNA-FISH identification could be $377.74/day. For coagulase-negative staphylococcus (CoNS, discontinuation of vancomycin could result in savings of $20.00/day. Conclusions In this retrospective study, excellent accuracy of PNA-FISH in blood and peritoneal fluids with reduced time to identification was observed, as compared to

  15. VIP Barcoding: composition vector-based software for rapid species identification based on DNA barcoding.

    Science.gov (United States)

    Fan, Long; Hui, Jerome H L; Yu, Zu Guo; Chu, Ka Hou

    2014-07-01

    Species identification based on short sequences of DNA markers, that is, DNA barcoding, has emerged as an integral part of modern taxonomy. However, software for the analysis of large and multilocus barcoding data sets is scarce. The Basic Local Alignment Search Tool (BLAST) is currently the fastest tool capable of handling large databases (e.g. >5000 sequences), but its accuracy is a concern and has been criticized for its local optimization. However, current more accurate software requires sequence alignment or complex calculations, which are time-consuming when dealing with large data sets during data preprocessing or during the search stage. Therefore, it is imperative to develop a practical program for both accurate and scalable species identification for DNA barcoding. In this context, we present VIP Barcoding: a user-friendly software in graphical user interface for rapid DNA barcoding. It adopts a hybrid, two-stage algorithm. First, an alignment-free composition vector (CV) method is utilized to reduce searching space by screening a reference database. The alignment-based K2P distance nearest-neighbour method is then employed to analyse the smaller data set generated in the first stage. In comparison with other software, we demonstrate that VIP Barcoding has (i) higher accuracy than Blastn and several alignment-free methods and (ii) higher scalability than alignment-based distance methods and character-based methods. These results suggest that this platform is able to deal with both large-scale and multilocus barcoding data with accuracy and can contribute to DNA barcoding for modern taxonomy. VIP Barcoding is free and available at http://msl.sls.cuhk.edu.hk/vipbarcoding/. © 2014 John Wiley & Sons Ltd.

  16. Rapid and accurate identification of Xanthomonas citri subspecies citri by fluorescence in situ hybridization.

    Science.gov (United States)

    Waite, D W; Griffin, R; Taylor, R; George, S

    2016-11-01

    Citrus canker is an economically important disease caused by the bacterial pathogen Xanthomonas citri subsp. citri (Xcc). This organism targets a wide range of citrus plants, including sweet orange, grapefruit, lemon and lime. As Xcc is spread by environmental factors such as wind and rain, it is difficult to control its movement once the disease has established. In order to facilitate monitoring of citrus canker we sought to design a novel diagnostic protocol based on fluorescence in situ hybridization (FISH) for identification of bacterial cells directly from canker pustules without cultivation or DNA extraction. This method was validated for specificity against a range of Xanthomonas species and strains. We show that our assay is extremely rapid (typically requiring between 2 and 3 h), and possesses a similar specificity to existing PCR diagnostic tools. The sensitivity of the assay is comparable to that of an existing PCR-based technique and sufficient for identifying Xcc in symptomatic plant material. The method is easily transferable to diagnosticians without prior experience using FISH. Xanthomonas citri subsp. citri (Xcc) is an aggressive and hardy pathogen of citrus plants worldwide. Outbreaks are difficult and costly to contain and the establishment of citrus canker results in restricted trade. In order to extend the existing toolkit for identification of Xcc we developed a novel diagnostic approach based on fluorescence in situ hybridization. Our approach is of comparable specificity and sensitivity to existing methods but can be performed directly on infected tissue making it significantly faster than existing PCRs, and requiring fewer laboratory resources. © 2016 The Society for Applied Microbiology.

  17. Improved method for rapid and accurate isolation and identification of Streptococcus mutans and Streptococcus sobrinus from human plaque samples.

    Science.gov (United States)

    Villhauer, Alissa L; Lynch, David J; Drake, David R

    2017-08-01

    Mutans streptococci (MS), specifically Streptococcus mutans (SM) and Streptococcus sobrinus (SS), are bacterial species frequently targeted for investigation due to their role in the etiology of dental caries. Differentiation of S. mutans and S. sobrinus is an essential part of exploring the role of these organisms in disease progression and the impact of the presence of either/both on a subject's caries experience. Of vital importance to the study of these organisms is an identification protocol that allows us to distinguish between the two species in an easy, accurate, and timely manner. While conducting a 5-year birth cohort study in a Northern Plains American Indian tribe, the need for a more rapid procedure for isolating and identifying high volumes of MS was recognized. We report here on the development of an accurate and rapid method for MS identification. Accuracy, ease of use, and material and time requirements for morphological differentiation on selective agar, biochemical tests, and various combinations of PCR primers were compared. The final protocol included preliminary identification based on colony morphology followed by PCR confirmation of species identification using primers targeting regions of the glucosyltransferase (gtf) genes of SM and SS. This method of isolation and identification was found to be highly accurate, more rapid than the previous methodology used, and easily learned. It resulted in more efficient use of both time and material resources. Copyright © 2017 Elsevier B.V. All rights reserved.

  18. Development of a multiplex PCR assay for rapid identification of Burkholderia pseudomallei, Burkholderia thailandensis, Burkholderia mallei and Burkholderia cepacia complex.

    Science.gov (United States)

    Koh, Seng Fook; Tay, Sun Tee; Sermswan, Rasana; Wongratanacheewin, Surasakdi; Chua, Kek Heng; Puthucheary, Savithri D

    2012-09-01

    We have developed a multiplex PCR assay for rapid identification and differentiation of cultures for Burkholderia pseudomallei, Burkholderia thailandensis, Burkholderia mallei and Burkholderia cepacia complex. The assay is valuable for use in clinical and veterinary laboratories, and in a deployable laboratory during outbreaks. Copyright © 2012 Elsevier B.V. All rights reserved.

  19. Pyrosequencing for rapid molecular identification of Schistosoma japonicum and S. mekongi eggs and cercariae.

    Science.gov (United States)

    Thanchomnang, Tongjit; Tantrawatpan, Chairat; Intapan, Pewpan M; Sri-Aroon, Pusadee; Limpanont, Yanin; Lulitanond, Viraphong; Janwan, Penchom; Sanpool, Oranuch; Tourtip, Somjintana; Maleewong, Wanchai

    2013-09-01

    Schistosomiasis, which is caused by Schistosoma japonicum and S. mekongi, is a chronic and dangerous widespread disease affecting several countries in Asia. Differentiation between S. japonicum and S. mekongi eggs and/or cercariae via microscopic examination is difficult due to morphological similarities. It is important to identify these etiological agents isolated from animals and humans at the species or genotype level. In this study, a pyrosequencing assay designed to detect S. japonicum and S. mekongi DNA in fecal samples and infected snails was developed and evaluated as an alternative tool to diagnose schistosomiasis. New primers targeting the 18S ribosomal RNA gene were designated for specific amplification. S. japonicum and S. mekongi were identified using a 43-nucleotide pattern of the 18S ribosomal RNA gene and were differentiated using 7 nucleotides within this region. S. japonicum and S. mekongi-infected snails and fecal samples derived from infected mice and rats were differentially detected within a short period of time. The analytical sensitivity of the method enabled the identification of as little as a single cercaria artificially introduced into a pool of 10 non-infected snails and 2 eggs inoculated in 100mg of non-infected fecal sample. To evaluate the comparative efficacy of the assay, identical samples were also analyzed via microscopy and Sanger sequencing. The pyrosequencing technique was found to be superior to the microscopy method and more rapid than the Sanger sequencing method. These results suggest that the pyrosequencing assay is rapid, simple, sensitive and accurate in identifying S. japonicum and S. mekongi in intermediate hosts and fecal samples of the final host. Copyright © 2013 Elsevier Inc. All rights reserved.

  20. Interaction between Rivers and Aquifers: a method for rapid Identification of Transience in Streambed Conductance

    Science.gov (United States)

    Gianni, Guillaume; Perrochet, Pierre; Vogel, Alexandre; Brunner, Philip

    2016-04-01

    Streambed hydraulic conductance controls the interactions between surface water and groundwater. In order to quantify river-aquifer dynamics, quantifying conductance is indispensable. However, the streambed conductance is often subject to transience, as a result of the erosion and deposition processes in rivers. This transience has to be quantified and considered for any approach (i.e. numerical or analytical models) aimed at quantifying exchange fluxes. Directly measuring hydraulic properties in a river yields only point values, is time-consuming and therefore not suited to detect transience of the physical properties. We present a method to continuously and rapidly monitor transience of streambed conductance. Input data are time series of stream stage and hydraulic head variations in the aquifer. The method is based on the inversion of a floodwave response. The analytical model consists of only 3 parameters: x, the distance between streambank and an observation well, α, the aquifer diffusivity and a, the retardation coefficient that is inversely proportional to the streambed conductance. Estimation of a is carried out over successive time steps in order to identify transience in streambed conductance. The method is tested on synthetic data and is applied to field data from the Rhône River and its alluvial aquifer (Switzerland). The synthetic method demonstrated the robustness of the proposed methodology. Application of the method to the field site allowed identifying transience in streambed properties, following flood events in the Rhône. This method requires transience in the surface water, and the river should not change its width significantly with a rising water level. If these conditions are fulfilled, this method allows for a rapid and effective identification of transience of streambed conductance.

  1. Field-deployable, quantitative, rapid identification of active Ebola virus infection in unprocessed blood.

    Science.gov (United States)

    Shah, Kavit; Bentley, Emma; Tyler, Adam; Richards, Kevin S R; Wright, Edward; Easterbrook, Linda; Lee, Diane; Cleaver, Claire; Usher, Louise; Burton, Jane E; Pitman, James K; Bruce, Christine B; Edge, David; Lee, Martin; Nazareth, Nelson; Norwood, David A; Moschos, Sterghios A

    2017-11-01

    The West African Ebola virus outbreak underlined the importance of delivering mass diagnostic capability outside the clinical or primary care setting in effectively containing public health emergencies caused by infectious disease. Yet, to date, there is no solution for reliably deploying at the point of need the gold standard diagnostic method, real time quantitative reverse transcription polymerase chain reaction (RT-qPCR), in a laboratory infrastructure-free manner. In this proof of principle work, we demonstrate direct performance of RT-qPCR on fresh blood using far-red fluorophores to resolve fluorogenic signal inhibition and controlled, rapid freeze/thawing to achieve viral genome extraction in a single reaction chamber assay. The resulting process is entirely free of manual or automated sample pre-processing, requires no microfluidics or magnetic/mechanical sample handling and thus utilizes low cost consumables. This enables a fast, laboratory infrastructure-free, minimal risk and simple standard operating procedure suited to frontline, field use. Developing this novel approach on recombinant bacteriophage and recombinant human immunodeficiency virus (HIV; Lentivirus), we demonstrate clinical utility in symptomatic EBOV patient screening using live, infectious Filoviruses and surrogate patient samples. Moreover, we evidence assay co-linearity independent of viral particle structure that may enable viral load quantification through pre-calibration, with no loss of specificity across an 8 log-linear maximum dynamic range. The resulting quantitative rapid identification (QuRapID) molecular diagnostic platform, openly accessible for assay development, meets the requirements of resource-limited countries and provides a fast response solution for mass public health screening against emerging biosecurity threats.

  2. A rapid and reliable species-specific identification of clinical and environmental isolates of Vibrio cholerae using a three-test procedure and recA polymerase chain reaction

    Directory of Open Access Journals (Sweden)

    A D Roozbehani

    2012-01-01

    Full Text Available Purpose: Vibrio cholerae, the cause of cholera, is one of the leading causes of morbidity and mortality in many developing countries. Most laboratories initially rely on biochemical tests for a presumptive identification of these strains, followed by a polymerase chain reaction (PCR-based method to confirm their identification. The aim of this study is to establish a rapid and reliable identification scheme for V. cholerae using a minimal, but highly specific number of biochemical tests and a PCR assay. Materials and Methods: We developed a species-specific PCR to identify V. cholerae, using a housekeeping gene recA, and used that to evaluate the sensitivity and specificity of 12 biochemical tests commonly used for screening and / or presumptive identification of V. cholerae in the clinical and environmental samples. Results: Here we introduced a combination of three biochemical tests, namely, sucrose fermentation, oxidase test, and growth in trypton broth containing 0% NaCl, as also the PCR of the recA gene, for rapid identification of V. cholerae isolates, with 100% sensitivity and specificity. The established method accurately identified a collection of 47 V. cholerae strains isolated from the clinical cases (n = 26 and surface waters (n = 21, while none of the 32 control strains belonging to different species were positive in this assay. Conclusion: The triple-test procedure introduced here is a simple and useful assay which can be adopted in cholera surveillance programs for efficient monitoring of V. cholerae in surface water and fecal samples.

  3. Integrated Biosensor Assay for Rapid Uropathogen Identification and Phenotypic Antimicrobial Susceptibility Testing.

    Science.gov (United States)

    Altobelli, Emanuela; Mohan, Ruchika; Mach, Kathleen E; Sin, Mandy Lai Yi; Anikst, Victoria; Buscarini, Maurizio; Wong, Pak Kin; Gau, Vincent; Banaei, Niaz; Liao, Joseph C

    2017-04-01

    Standard diagnosis of urinary tract infection (UTI) via urine culture for pathogen identification (ID) and antimicrobial susceptibility testing (AST) takes 2-3 d. This delay results in empiric treatment and contributes to the misuse of antibiotics and the rise of resistant pathogens. A rapid diagnostic test for UTI may improve patient care and antibiotic stewardship. To develop and validate an integrated biosensor assay for UTI diagnosis, including pathogen ID and AST, with determination of the minimum inhibitory concentration (MIC) for ciprofloxacin. Urine samples positive for Enterobacteriaceae (n=84) or culture-negative (n=23) were obtained from the Stanford Clinical Microbiology Laboratory between November 2013 and September 2014. Each sample was diluted and cultured for 5h with and without ciprofloxacin, followed by quantitative detection of bacterial 16S rRNA using a single electrochemical biosensor array functionalized with a panel of complementary DNA probes. Pathogen ID was determined using universal bacterial, Enterobacteriaceae (EB), and pathogen-specific probes. Phenotypic AST with ciprofloxacin MIC was determined using an EB probe to measure 16S rRNA levels as a function of bacterial growth. Electrochemical signals for pathogen ID at 6 SD over background were considered positive. An MIC signal of 0.4 log units lower than the no-antibiotic control indicated sensitivity. Results were compared to clinical microbiology reports. For pathogen ID, the assay had 98.5% sensitivity, 96.6% specificity, 93.0% positive predictive value, and 99.3% negative predictive value. For ciprofloxacin MIC the categorical and essential agreement was 97.6%. Further automation, testing of additional pathogens and antibiotics, and a full prospective study will be necessary for translation to clinical use. The integrated biosensor platform achieved microbiological results including MIC comparable to standard culture in a significantly shorter assay time. Further assay automation

  4. Real time PCR for the rapid identification and drug susceptibility of Mycobacteria present in Bronchial washings

    Directory of Open Access Journals (Sweden)

    Thilini Piushani Keerthirathne

    2016-10-01

    Full Text Available Abstract Background Mycobacteria have a spectrum of virulence and different susceptibilities to antibiotics. Distinguishing mycobacterial species is vital as patients with non-tuberculous mycobacterial (NTM infections present clinical features that are similar to those of patients with tuberculosis. Thus, rapid differentiation of Mycobacterium tuberculosis complex from NTM is critical to administer appropriate treatment. Hence the aim of the study was to rapid identification of mycobacterial species present in bronchial washings using multiplex real time Polymerase Chain Reaction (PCR and to determine the drug susceptibility in identified mycobacterial species. Methods Sputum smear negative bronchoscopy specimens (n = 150 were collected for a period of one year, from patients attending the General Hospital Kandy, Sri Lanka. The specimens were processed with modified Petroff’s method and were cultured on Löwenstein– Jensen medium. DNA, extracted from the mycobacterial isolates were subjected to a SYBR green mediated real time multiplex, PCR assay with primers specific for the M. tuberculosis complex, M. avium complex, M. chelonae-M.abscessus group and M. fortuitum group. DNA sequencing was performed for the species confirmation, by targeting the 16S rRNA gene and the drug susceptibility testing was performed for the molecularly identified isolates of M. tuberculosis and NTM. Results The optimized SYBR Green mediated multiplex real-time PCR assay was able to identify the presence of genus Mycobacterium in 25 out of 26 AFB positive isolates, two M. tuberculosis complex, three M. avium complex and two isolates belonging to M. chelonae-M. abscessus group. DNA sequencing confirmed the presence of M. tuberculosis, M. chelonae-M. abscessus, M. intracellulare, M. avium, Rhodococcus sp. and M. celatum. Remaining isolates were identified as Mycobacterium sp. All the NTM isolates were sensitive to amikacin and seven were resistant to ciproflaxacin

  5. Two new media Pinus halepensis seed agar and blackberry agar for rapid identification of Cryptococcus neoformans.

    Science.gov (United States)

    Mseddi, Fatma; Sellami, Amira; Sellami, Hayet; Cheikhrouhou, Fatma; Makni, Fattouma; Ayadi, Ali

    2011-07-01

    Cryptococcus neoformans is an encapsulated yeast-like fungus that causes life-threatening infections, particularly in immunocompromised patients. The formation of brown pigment on many media described in the literature, such as that in Niger seed (Guizotia abyssinica) agar, has been used to identify C. neoformans. The present study compares melanin production by clinical and environmental isolates of C. neoformans and other medically important yeast on two new media, Pinus halepensis seed (PHS) agar and blackberry (BlaB) agar, and the classic medium Niger seed agar. Results obtained after the culture of 46 strains of C. neoformans, for 4, 24 and 48 h at 37 °C on these three media, showed that at 24 h, 100% of strains were pigmented on BlaB agar, 91.3% on PHS agar but only 34.8% on Niger seed agar. In conclusion, PHS and BlaB agar are two interesting new media for the rapid identification of C. neoformans isolates. © 2010 Blackwell Verlag GmbH.

  6. An integrated lab-on-chip for rapid identification and simultaneous differentiation of tropical pathogens.

    Science.gov (United States)

    Tan, Jeslin J L; Capozzoli, Monica; Sato, Mitsuharu; Watthanaworawit, Wanitda; Ling, Clare L; Mauduit, Marjorie; Malleret, Benoît; Grüner, Anne-Charlotte; Tan, Rosemary; Nosten, François H; Snounou, Georges; Rénia, Laurent; Ng, Lisa F P

    2014-01-01

    Tropical pathogens often cause febrile illnesses in humans and are responsible for considerable morbidity and mortality. The similarities in clinical symptoms provoked by these pathogens make diagnosis difficult. Thus, early, rapid and accurate diagnosis will be crucial in patient management and in the control of these diseases. In this study, a microfluidic lab-on-chip integrating multiplex molecular amplification and DNA microarray hybridization was developed for simultaneous detection and species differentiation of 26 globally important tropical pathogens. The analytical performance of the lab-on-chip for each pathogen ranged from 102 to 103 DNA or RNA copies. Assay performance was further verified with human whole blood spiked with Plasmodium falciparum and Chikungunya virus that yielded a range of detection from 200 to 4×105 parasites, and from 250 to 4×107 PFU respectively. This lab-on-chip was subsequently assessed and evaluated using 170 retrospective patient specimens in Singapore and Thailand. The lab-on-chip had a detection sensitivity of 83.1% and a specificity of 100% for P. falciparum; a sensitivity of 91.3% and a specificity of 99.3% for P. vivax; a positive 90.0% agreement and a specificity of 100% for Chikungunya virus; and a positive 85.0% agreement and a specificity of 100% for Dengue virus serotype 3 with reference methods conducted on the samples. Results suggested the practicality of an amplification microarray-based approach in a field setting for high-throughput detection and identification of tropical pathogens.

  7. Rapid identification of paragonimiasis foci by lay informants in Lao People's Democratic Republic.

    Directory of Open Access Journals (Sweden)

    Peter Odermatt

    Full Text Available BACKGROUND: Paragonimiasis is a food-borne trematodiasis leading to lung disease. Worldwide, an estimated 21 million people are infected. Foci of ongoing transmission remain often unnoticed. We evaluated a simple questionnaire approach using lay-informants at the village level to identify paragonimiasis foci and suspected paragonimiasis cases. METHODOLOGY/PRINCIPAL FINDINGS: The study was carried out in an endemic area of Lao People's Democratic Republic. Leaders of 49 remote villages in northern Vientiane Province were asked to notify suspected paragonimiasis patients using a four-item questionnaire sent through administrative channels: persons responding positively for having chronic cough (more than 3 weeks and/or blood in sputum with or without fever. We validated the village leaders' reports in ten representative villages with a door-to-door survey. We examined three sputa of suspected patients for the presence of Paragonimus eggs and acid fast bacilli. 91.8% of village leaders participated and notified a total of 220 suspected patients; 76.2% were eventually confirmed; an additional 138 suspected cases were found in the survey. Sensitivity of village leaders' notice for "chronic cough" and "blood in sputum" was 100%; "blood in sputum" alone reached a sensitivity of 85.7%. SIGNIFICANCE: Our approach led to the identification of three previously unknown foci of transmission. A rapid and simple lay-informant questionnaire approach is a promising low-cost community diagnostic tool of paragonimiasis control programs.

  8. Rapid identification of transience in streambed conductance by inversion of floodwave responses

    Science.gov (United States)

    Gianni, Guillaume; Richon, Julien; Perrochet, Pierre; Vogel, Alexandre; Brunner, Philip

    2016-04-01

    Streambed conductance controls the interaction between surface and groundwater. However, the streambed conductance is often subject to transience. Directly measuring hydraulic properties in a river yields only point values, is time-consuming and therefore not suited to detect transience of physical properties. Here, we present a method to continuously monitor transience in streambed conductance. Input data are time series of stream stage and near stream hydraulic head. The method is based on the inversion of floodwave responses. The analytical model consists of three parameters: x, the distance between streambank and an observation well, α, the aquifer diffusivity, and a the retardation coefficient that is inversely proportional to the streambed conductance. Estimation of a is carried out over successive time steps in order to identify transience in streambed conductance. The method is tested using synthetic data and is applied to field data from the Rhône River and its alluvial aquifer (Switzerland). The synthetic method demonstrated the robustness of the proposed methodology. Application of the method to the field data allowed identifying transience in streambed properties, following flood events in the Rhône. This method requires transience in the surface water, and the river should not change its width significantly with a rising water level. If these conditions are fulfilled, this method allows for a rapid and effective identification of transience in streambed conductance.

  9. Portable platform for rapid in-field identification of human fecal pollution in water.

    Science.gov (United States)

    Jiang, Yu Sherry; Riedel, Timothy E; Popoola, Jessica A; Morrow, Barrett R; Cai, Sheng; Ellington, Andrew D; Bhadra, Sanchita

    2017-12-13

    Human fecal contamination of water is a public health risk. However, inadequate testing solutions frustrate timely, actionable monitoring. Bacterial culture-based methods are simple but typically cannot distinguish fecal host source. PCR assays can identify host sources but require expertise and infrastructure. To bridge this gap we have developed a field-ready nucleic acid diagnostic platform and rapid sample preparation methods that enable on-site identification of human fecal contamination within 80 min of sampling. Our platform relies on loop-mediated isothermal amplification (LAMP) of human-associated Bacteroides HF183 genetic markers from crude samples. Oligonucleotide strand exchange (OSD) probes reduce false positives by sequence specifically transducing LAMP amplicons into visible fluorescence that can be photographed by unmodified smartphones. Our assay can detect as few as 17 copies/ml of human-associated HF183 targets in sewage-contaminated water without cross-reaction with canine or feline feces. It performs robustly with a variety of environmental water sources and with raw sewage. We have also developed lyophilized assays and inexpensive 3D-printed devices to minimize cost and facilitate field application. Copyright © 2017 Elsevier Ltd. All rights reserved.

  10. Rapid Detection and Identification of Infectious Pathogens Based on High-throughput Sequencing

    Directory of Open Access Journals (Sweden)

    Pei-Xiang Ni

    2015-01-01

    Full Text Available Background: The dilemma of pathogens identification in patients with unidentified clinical symptoms such as fever of unknown origin exists, which not only poses a challenge to both the diagnostic and therapeutic process by itself, but also to expert physicians. Methods: In this report, we have attempted to increase the awareness of unidentified pathogens by developing a method to investigate hitherto unidentified infectious pathogens based on unbiased high-throughput sequencing. Results: Our observations show that this method supplements current diagnostic technology that predominantly relies on information derived five cases from the intensive care unit. This methodological approach detects viruses and corrects the incidence of false positive detection rates of pathogens in a much shorter period. Through our method is followed by polymerase chain reaction validation, we could identify infection with Epstein-Barr virus, and in another case, we could identify infection with Streptococcus viridians based on the culture, which was false positive. Conclusions: This technology is a promising approach to revolutionize rapid diagnosis of infectious pathogens and to guide therapy that might result in the improvement of personalized medicine.

  11. 1-1-12 one-step wash-in scheme for desflurane-nitrous oxide low-flow anesthesia: rapid and predictable induction.

    Science.gov (United States)

    Sathitkarnmanee, Thepakorn; Tribuddharat, Sirirat; Suttinarakorn, Chakthip; Nonlhaopol, Duangthida; Thananun, Maneerat; Somdee, Wilawan; Theerapongpakdee, Sunchai

    2014-01-01

    We propose a 1-1-12 wash-in scheme for desflurane-nitrous oxide (N2O) low-flow anesthesia. The objective of our study was to determine the time to achieve alveolar concentration of desflurane (FAD) at 1, 2, 3, 4, 5, and 6%. We enrolled 106 patients scheduled for elective surgery under general anesthesia. After induction and intubation, wash-in was started with a fresh gas flow (FGF) of N2O : O2 1 : 1 L min(-1) and vaporizer concentration of desflurane (FD) of 12%. Ventilation was controlled to maintain PACO2 at 30-35 mmHg. The FAD rose rapidly from 0 to 4% in 2 min in a linear manner in 0.5 min increments. An FAD of 6% was achieved in 4 min in a linear fashion from FAD of 4% but in 1 min increments. An FAD of 1 to 6% occurred at 0.6, 1, 1.5, 2, 3, and 4 min. Heart rate during wash-in showed a statistically, albeit not clinically, significant pattern of increase. By contrast, blood pressure slightly decreased during this period. We developed a 1-1-12 wash-in scheme using a FGF of N2O : O2 1 : 1 L min(-1) and FD of 12% for desflurane-nitrous oxide low-flow anesthesia. A respective FAD of 1, 2, 3, 4, 5, and 6% can be expected at 0.6, 1, 1.5, 2, 3, and 4 min.

  12. 1-1-12 One-Step Wash-In Scheme for Desflurane-Nitrous Oxide Low-Flow Anesthesia: Rapid and Predictable Induction

    Directory of Open Access Journals (Sweden)

    Thepakorn Sathitkarnmanee

    2014-01-01

    Full Text Available Background. We propose a 1-1-12 wash-in scheme for desflurane-nitrous oxide (N2O low-flow anesthesia. The objective of our study was to determine the time to achieve alveolar concentration of desflurane (FAD at 1, 2, 3, 4, 5, and 6%. Methods. We enrolled 106 patients scheduled for elective surgery under general anesthesia. After induction and intubation, wash-in was started with a fresh gas flow (FGF of N2O : O2 1 : 1 L min−1 and vaporizer concentration of desflurane (FD of 12%. Ventilation was controlled to maintain PACO2 at 30–35 mmHg. Results. The FAD rose rapidly from 0 to 4% in 2 min in a linear manner in 0.5 min increments. An FAD of 6% was achieved in 4 min in a linear fashion from FAD of 4% but in 1 min increments. An FAD of 1 to 6% occurred at 0.6, 1, 1.5, 2, 3, and 4 min. Heart rate during wash-in showed a statistically, albeit not clinically, significant pattern of increase. By contrast, blood pressure slightly decreased during this period. Conclusions. We developed a 1-1-12 wash-in scheme using a FGF of N2O : O2 1 : 1 L min−1 and FD of 12% for desflurane-nitrous oxide low-flow anesthesia. A respective FAD of 1, 2, 3, 4, 5, and 6% can be expected at 0.6, 1, 1.5, 2, 3, and 4 min.

  13. Development of a novel, simple and rapid molecular identification system for clinical Candida species

    NARCIS (Netherlands)

    Deak, R.; Bodia, L.; Aarts, H.J.M.; Maraz, A.

    2004-01-01

    Identification of clinical yeast isolates causing candidiasis is routinely performed by commercial yeast identification systems based on biochemical, morphological and physiological tests. These systems require 3-5 days and the proportion of identifications that are incorrect is high. Our novel and

  14. Application of MALDI-TOF MS Systems in the Rapid Identification of Campylobacter spp. of Public Health Importance.

    Science.gov (United States)

    Hsieh, Ying-Hsin; Wang, Yun F; Moura, Hercules; Miranda, Nancy; Simpson, Steven; Gowrishankar, Ramnath; Barr, John; Kerdahi, Khalil; Sulaiman, Irshad M

    2017-09-12

    Campylobacteriosis is an infectious gastrointestinal disease caused by Campylobacter spp.In most cases, it is either underdiagnosed or underreported due to poor diagnostics and limited databases. Several DNA-based molecular diagnostic techniques, including 16S ribosomal RNA (rRNA) sequence typing, have been widely used in the species identification of Campylobacter. Nevertheless, these assays are time-consuming and require a high quality of bacterial DNA. Matrix-assisted laser desorption ionization (MALDI) time-of-flight (TOF) MS is an emerging diagnostic technology that can provide the rapid identification of microorganisms by using their intact cells without extraction or purification. In this study, we analyzed 24 American Type Culture Collection reference isolates of 16 Campylobacter spp. and five unknown clinical bacterial isolates for rapid identification utilizing two commercially available MADI-TOF MS platforms, namely the bioMérieux VITEK(®) MS and Bruker Biotyper systems. In addition, 16S rRNA sequencing was performed to confirm the species-level identification of the unknown clinical isolates. Both MALDI-TOF MS systems identified the isolates of C. jejuni, C. coli, C. lari, and C. fetus. The results of this study suggest that the MALDI-TOF MS technique can be used in the identification of Campylobacter spp. of public health importance.

  15. A robust haze-removal scheme in polarimetric dehazing imaging based on automatic identification of sky region

    Science.gov (United States)

    Zhang, Wenfei; Liang, Jian; Ju, Haijuan; Ren, Liyong; Qu, Enshi; Wu, Zhaoxin

    2016-12-01

    Quality enhancement of images acquired in hazy conditions is a significant research area in civil and military applications. The polarimetric dehazing methods have been exploited to dehaze hazy images and have proven to be effective in enhancing their quality. In this paper, by combining the polarimetric imaging technique and the dark channel prior technique, a robust haze-removal scheme is presented for the first time. On the one hand, the polarimetric imaging technique has advantages in recovering detailed information well, especially in dense hazy conditions; on the other hand, the dark channel prior technique provides a much more precise and convenient way to estimate the airlight radiance through extracting the sky region automatically. The experiments verify the practicability and effectiveness of the proposed dehazing scheme in quality enhancement of hazy images. Furthermore, comparison study demonstrates that the proposed scheme is superior to some sophisticated methods in terms of the visibility and contrast. We believe this scheme is beneficial in the image dehazing applications, especially for real-time applications.

  16. Identification of "Streptococcus milleri" group isolates to the species level with a commercially available rapid test system.

    Science.gov (United States)

    Flynn, C E; Ruoff, K L

    1995-01-01

    Clinical isolates of the "Streptococcus milleri" species group were examined by conventional methods and a rapid, commercially available method for the identification of these strains to the species level. The levels of agreement between the identifications obtained with the commercially available system (Fluo-Card Milleri; KEY Scientific, Round Rock, Tex.) and conventional methods were 98% for 50 Streptococcus anginosus strains, 97% for 31 Streptococcus constellatus strains, and 88% for 17 isolates identified as Streptococcus intermedius. Patient records were also studied in order to gain information on the frequency and sites of isolation of each of the three "S. milleri" group species. PMID:8567909

  17. Rapid identification of Pterocarpus santalinus and Dalbergia louvelii by FTIR and 2D correlation IR spectroscopy

    Science.gov (United States)

    Zhang, Fang-Da; Xu, Chang-Hua; Li, Ming-Yu; Huang, An-Min; Sun, Su-Qin

    2014-07-01

    Since Pterocarpus santalinus and Dalbergia louvelii, which are of precious Rosewood, are very similar in their appearance and anatomy characteristics, cheaper Hongmu D. louvelii is often illegally used to impersonate valuable P. santalinus, especially in Chinese furniture manufacture. In order to develop a rapid and effective method for easy confused wood furniture differentiation, we applied tri-step identification method, i.e., conventional infrared spectroscopy (FT-IR), second derivative infrared (SD-IR) spectroscopy and two-dimensional correlation infrared (2DCOS-IR) spectroscopy to investigate P. santalinus and D. louvelii furniture. According to FT-IR and SD-IR spectra, it has been found two unconditional stable difference at 848 cm-1 and 700 cm-1 and relative stable differences at 1735 cm-1, 1623 cm-1, 1614 cm-1, 1602 cm-1, 1509 cm-1, 1456 cm-1, 1200 cm-1, 1158 cm-1, 1055 cm-1, 1034 cm-1 and 895 cm-1 between D. louvelii and P. santalinus IR spectra. The stable discrepancy indicates that the category of extractives is different between the two species. Besides, the relative stable differences imply that the content of holocellulose in P. santalinus is more than that of D. louvelii, whereas the quantity of extractives in D. louvelii is higher. Furthermore, evident differences have been observed in their 2DCOS-IR spectra of 1550-1415 cm-1 and 1325-1030 cm-1. P. santalinus has two strong auto-peaks at 1459 cm-1 and 1467 cm-1, three mid-strong auto-peaks at 1518 cm-1, 1089 cm-1 and 1100 cm-1 and five weak auto-peaks at 1432 cm-1, 1437 cm-1, 1046 cm-1, 1056 cm-1 and 1307 cm-1 while D. louvelii has four strong auto-peaks at 1465 cm-1, 1523 cm-1, 1084 cm-1 and 1100 cm-1, four mid-strong auto-peaks at 1430 cm-1, 1499 cm-1, 1505 cm-1 and 1056 cm-1 and two auto-peaks at 1540 cm-1 and 1284 cm-1. This study has proved that FT-IR integrated with 2DCOS-IR could be applicable for precious wood furniture authentication in a direct, rapid and holistic manner.

  18. An Integrated Lab-on-Chip for Rapid Identification and Simultaneous Differentiation of Tropical Pathogens

    Science.gov (United States)

    Sato, Mitsuharu; Watthanaworawit, Wanitda; Ling, Clare L.; Mauduit, Marjorie; Malleret, Benoît; Grüner, Anne-Charlotte; Tan, Rosemary; Nosten, François H.; Snounou, Georges; Rénia, Laurent; Ng, Lisa F. P.

    2014-01-01

    Tropical pathogens often cause febrile illnesses in humans and are responsible for considerable morbidity and mortality. The similarities in clinical symptoms provoked by these pathogens make diagnosis difficult. Thus, early, rapid and accurate diagnosis will be crucial in patient management and in the control of these diseases. In this study, a microfluidic lab-on-chip integrating multiplex molecular amplification and DNA microarray hybridization was developed for simultaneous detection and species differentiation of 26 globally important tropical pathogens. The analytical performance of the lab-on-chip for each pathogen ranged from 102 to 103 DNA or RNA copies. Assay performance was further verified with human whole blood spiked with Plasmodium falciparum and Chikungunya virus that yielded a range of detection from 200 to 4×105 parasites, and from 250 to 4×107 PFU respectively. This lab-on-chip was subsequently assessed and evaluated using 170 retrospective patient specimens in Singapore and Thailand. The lab-on-chip had a detection sensitivity of 83.1% and a specificity of 100% for P. falciparum; a sensitivity of 91.3% and a specificity of 99.3% for P. vivax; a positive 90.0% agreement and a specificity of 100% for Chikungunya virus; and a positive 85.0% agreement and a specificity of 100% for Dengue virus serotype 3 with reference methods conducted on the samples. Results suggested the practicality of an amplification microarray-based approach in a field setting for high-throughput detection and identification of tropical pathogens. PMID:25078474

  19. An integrated lab-on-chip for rapid identification and simultaneous differentiation of tropical pathogens.

    Directory of Open Access Journals (Sweden)

    Jeslin J L Tan

    Full Text Available Tropical pathogens often cause febrile illnesses in humans and are responsible for considerable morbidity and mortality. The similarities in clinical symptoms provoked by these pathogens make diagnosis difficult. Thus, early, rapid and accurate diagnosis will be crucial in patient management and in the control of these diseases. In this study, a microfluidic lab-on-chip integrating multiplex molecular amplification and DNA microarray hybridization was developed for simultaneous detection and species differentiation of 26 globally important tropical pathogens. The analytical performance of the lab-on-chip for each pathogen ranged from 102 to 103 DNA or RNA copies. Assay performance was further verified with human whole blood spiked with Plasmodium falciparum and Chikungunya virus that yielded a range of detection from 200 to 4×105 parasites, and from 250 to 4×107 PFU respectively. This lab-on-chip was subsequently assessed and evaluated using 170 retrospective patient specimens in Singapore and Thailand. The lab-on-chip had a detection sensitivity of 83.1% and a specificity of 100% for P. falciparum; a sensitivity of 91.3% and a specificity of 99.3% for P. vivax; a positive 90.0% agreement and a specificity of 100% for Chikungunya virus; and a positive 85.0% agreement and a specificity of 100% for Dengue virus serotype 3 with reference methods conducted on the samples. Results suggested the practicality of an amplification microarray-based approach in a field setting for high-throughput detection and identification of tropical pathogens.

  20. Rapid identification of novel immunodominant proteins and characterization of a specific linear epitope of Campylobacter jejuni.

    Directory of Open Access Journals (Sweden)

    Sebastian Hoppe

    Full Text Available Campylobacter jejuni remains one of the major gut pathogens of our time. Its zoonotic nature and wide-spread distribution in industrialized countries calls for a quick and reliable diagnostic tool. Antibody-based detection presents a suitable means to identify pathogenic bacteria. However, the knowledge about immunodominant targets is limited. Thus, an approach is presented, which allows for the rapid screening of numerous cDNA derived expression clones to identify novel antigens. The deeper understanding of immunodominant proteins assists in the design of diagnostic tools and furthers the insight into the bacterium's pathogenicity as well as revealing potential candidates for vaccination. We have successfully screened 1536 clones of an expression library to identify 22 proteins that have not been described as immunodominant before. After subcloning the corresponding 22 genes and expression of full-length proteins, we investigated the immunodominant character by microarrays and ELISA. Subsequently, seven proteins were selected for epitope mapping. For cj0669 and cj0920c linear epitopes were identified. For cj0669, specificity assays revealed a specific linear epitope site. Consequently, an eleven amino acid residue sequence TLIKELKRLGI was analyzed via alanine scan, which revealed the glycine residue to be significant for binding of the antibody. The innovative approach presented herein of generating cDNAs of prokaryotes in combination with a microarray platform rendering time-consuming purification steps obsolete has helped to illuminate novel immunodominant proteins of C.jejuni. The findings of a specific linear epitope pave the way for a plethora of future research and the potential use in diagnostic applications such as serological screenings. Moreover, the current approach is easily adaptable to other highly relevant bacteria making it a formidable tool for the future discovery of antigens and potential biomarkers. Consequently, it is

  1. Rapid detection and identification of Brachyspira aalborgi from rectal biopsies and faeces of a patient.

    Science.gov (United States)

    Calderaro, Adriana; Villanacci, Vincenzo; Conter, Mauro; Ragni, Patrizia; Piccolo, Giovanna; Zuelli, Claudia; Bommezzadri, Simona; Guégan, Rozenn; Zambelli, Claudia; Perandin, Francesca; Arcangeletti, Maria Cristina; Medici, Maria Cristina; Manca, Nino; Dettori, Giuseppe; Chezzi, Carlo

    2003-03-01

    This study reports for the first time the detection of Brachyspira aalborgi in faeces and rectal biopsies of a female suffering for 3-4 months of abdominal pain with long-standing mucosal diarrhoea, rectal bleeding and suspected carcinoma of the rectum. After pre-treatment of samples (faeces and biopsies) with a liquid medium (trypticase soy broth-TSB) containing foetal calf serum (FCS, 10%) and spectinomycin and rifampicin (TSB-SR) the first detection of B. aalborgi isolate HBS1 was observed after 48 h in the primary plates of selective blood agar modified medium (BAM) containing spectinomycin and rifampicin (BAM-SR), where growth zones were signalled by a small weakly beta-haemolytic halo. Attempts to subculture spirochaetes in agar media failed. The new HBS1 isolate was only propagated in TSB broth and at electron microscopy it showed 4 endoflagella inserted at each tapered end. The phenotypic characterization of HBS1 demonstrated absence of hippurate hydrolysis, indole production, alpha-galactosidase, alpha- and beta-glucosidase activities in accordance with the B. aalborgi type strain. Rapid identification of B. aalborgi isolate HBS1 was performed directly from faeces and rectal biopsies and subsequently from pure cultures by a genetic method based on 16S DNA restriction fragment length polymorphism (RFLP)-polymerase chain reaction (PCR). The sequence of 16S DNA amplicon of the isolate HBS1 was found 99.2% corresponding to that of the B. aalborgi type strain. Our results encourage further investigations for the development of a suitable selective agar medium for the isolating and cultivating B. aalborgi from human specimens.

  2. Rapid Identification of Pathogens from Positive Blood Cultures by Multiplex PCR using the FilmArray System

    Science.gov (United States)

    Blaschke, Anne J.; Heyrend, Caroline; Byington, Carrie L.; Fisher, Mark A.; Barker, Elizabeth; Garrone, Nicholas F.; Thatcher, Stephanie A.; Pavia, Andrew T.; Barney, Trenda; Alger, Garrison D.; Daly, Judy A.; Ririe, Kirk M.; Ota, Irene; Poritz, Mark A.

    2012-01-01

    Sepsis is a leading cause of death. Rapid and accurate identification of pathogens and antimicrobial resistance directly from blood culture could improve patient outcomes. The FilmArray® (FA; Idaho Technology, Inc., Salt Lake City, UT) Blood Culture (BC) panel can identify > 25 pathogens and 4 antibiotic resistance genes from positive blood cultures in 1 hour. We compared a development version of the panel to conventional culture and susceptibility testing on 102 archived blood cultures from adults and children with bacteremia. Of 109 pathogens identified by culture, 95% were identified by FA. Among 111 prospectively collected blood cultures, the FA identified 84 of 92 pathogens (91%) covered by the panel. Among 25 Staphylococcus aureus and 21 Enterococcus species detected, FA identified all culture-proven MRSA and VRE. The FA BC panel is an accurate method for the rapid identification of pathogens and resistance genes from blood culture. PMID:22999332

  3. Rapid disaster victim identification in mass fatality incidents: decision-support tool to facilitate human remains identification.

    Science.gov (United States)

    de Cosmo, Sergio; Barbera, Joseph A

    2012-10-01

    A quantitative decision-support tool (DST), using a combination of selected human physical attributes as identification elements, was developed to facilitate body identification in mass fatality incidents, particularly in settings with limited availability of technological resources and forensic expertise. To construct the DST, the external biological attributes of interest were first selected. A process was then developed to guide collection of the selected categories of attributes and record them into objective antemortem (AM) and postmortem (PM) records. Finally, a framework for assessing the similarity between confronting PM-AM attribute records was established. The DST evaluates the similarities between each set of like attributes in the AM and PM records being compared. It then computes an overall similarity score for each evaluated AM record that was compared to a selected PM record. The AM record with the highest score represents the highest probable match, with the PM file selected for the comparison. Multiple simulations across a range of mass fatality situations demonstrated the effectiveness of the DST in the experimental setting. The developed DST may provide authorities with a method for expediting body identification without completely eliminating any missing person file from consideration. Under specific circumstances, this method may reduce the need for technologically sophisticated forensic identification techniques (eg, dental records, fingerprints, and DNA). At a minimum, it should facilitate the efficiency of the current technological matching process.

  4. Evaluation of FilmArray and Verigene systems for rapid identification of positive blood cultures.

    Science.gov (United States)

    Bhatti, M M; Boonlayangoor, S; Beavis, K G; Tesic, V

    2014-09-01

    The Verigene tests for Gram-positive and Gram-negative organisms in blood culture and the FilmArray blood culture identification panel were assessed for their ability to identify pathogens from positive blood cultures. Both platforms correctly identified bacteria in 92% of monomicrobial cultures analyzed, with times to identification that were significantly shorter than those for identification from subcultures. Copyright © 2014, American Society for Microbiology. All Rights Reserved.

  5. Evaluation of the Rapid Mastitis Test for identification of Staphylococcus aureus and Streptococcus agalactiae isolated from bovine mammary glands.

    OpenAIRE

    Watts, J L; Owens, W E

    1988-01-01

    A latex agglutination test system (Rapid Mastitis Test [RMT]; Immucell, Portland, Maine) containing reagents for the identification of Staphylococcus aureus and Streptococcus agalactiae from bovine intramammary infections was evaluated with 527 staphylococcal and 267 streptococcal isolates. The RMT Staphylococcus aureus reagent detected 94.2% of 242 Staphylococcus aureus isolates, 80% of 25 Staphylococcus intermedius isolates, and 42.8% of 21 tube coagulase-positive Staphylococcus hyicus isol...

  6. The Department of Veterans Affairs Nutritional Status Classification Scheme Allows for Rapid Assessment of Nutritional Status Prior to Autologous Peripheral Blood Stem Cell Transplantation and Identifies Patients at High Risk of Transplant-Related Complications

    National Research Council Canada - National Science Library

    Toro, Juan J; Haile, David J; Chao, Ju-Hsien; Schneider, Deanna; Jewell, Pamela S; Lee, Shuko; Freytes, César O

    2009-01-01

    ... of Veterans Affairs (VA) developed a Nutritional Status Classification Scheme (NSCS) to identify nutritionally compromised inpatients rapidly and reliably. The VA-NSCS takes into account a combination of body weight, routine laboratory tests, and clinical and dietary history. The VA-NSCS is routinely utilized for the nutritional evaluation ...

  7. Rapid Molecular Identification of Pathogenic Yeasts by Pyrosequencing Analysis of 35 Nucleotides of Internal Transcribed Spacer 2 ▿

    Science.gov (United States)

    Borman, Andrew M.; Linton, Christopher J.; Oliver, Debra; Palmer, Michael D.; Szekely, Adrien; Johnson, Elizabeth M.

    2010-01-01

    Rapid identification of yeast species isolates from clinical samples is particularly important given their innately variable antifungal susceptibility profiles. Here, we have evaluated the utility of pyrosequencing analysis of a portion of the internal transcribed spacer 2 region (ITS2) for identification of pathogenic yeasts. A total of 477 clinical isolates encompassing 43 different fungal species were subjected to pyrosequencing analysis in a strictly blinded study. The molecular identifications produced by pyrosequencing were compared with those obtained using conventional biochemical tests (AUXACOLOR2) and following PCR amplification and sequencing of the D1-D2 portion of the nuclear 28S large rRNA gene. More than 98% (469/477) of isolates encompassing 40 of the 43 fungal species tested were correctly identified by pyrosequencing of only 35 bp of ITS2. Moreover, BLAST searches of the public synchronized databases with the ITS2 pyrosequencing signature sequences revealed that there was only minimal sequence redundancy in the ITS2 under analysis. In all cases, the pyrosequencing signature sequences were unique to the yeast species (or species complex) under investigation. Finally, when pyrosequencing was combined with the Whatman FTA paper technology for the rapid extraction of fungal genomic DNA, molecular identification could be accomplished within 6 h from the time of starting from pure cultures. PMID:20702674

  8. Rapid Evaporative Ionisation Mass Spectrometry (REIMS) Provides Accurate Direct from Culture Species Identification within the Genus Candida.

    Science.gov (United States)

    Cameron, Simon J S; Bolt, Frances; Perdones-Montero, Alvaro; Rickards, Tony; Hardiman, Kate; Abdolrasouli, Alireza; Burke, Adam; Bodai, Zsolt; Karancsi, Tamas; Simon, Daniel; Schaffer, Richard; Rebec, Monica; Balog, Julia; Takáts, Zoltan

    2016-11-14

    Members of the genus Candida, such as C. albicans and C. parapsilosis, are important human pathogens. Other members of this genus, previously believed to carry minimal disease risk, are increasingly recognised as important human pathogens, particularly because of variations in susceptibilities to widely used anti-fungal agents. Thus, rapid and accurate identification of clinical Candida isolates is fundamental in ensuring timely and effective treatments are delivered. Rapid Evaporative Ionisation Mass Spectrometry (REIMS) has previously been shown to provide a high-throughput platform for the rapid and accurate identification of bacterial and fungal isolates. In comparison to commercially available matrix assisted laser desorption ionisation time of flight mass spectrometry (MALDI-ToF), REIMS based methods require no preparative steps nor time-consuming cell extractions. Here, we report on the ability of REIMS-based analysis to rapidly and accurately identify 153 clinical Candida isolates to species level. Both handheld bipolar REIMS and high-throughput REIMS platforms showed high levels of species classification accuracy, with 96% and 100% of isolates classified correctly to species level respectively. In addition, significantly different (FDR corrected P value < 0.05) lipids within the 600 to 1000 m/z mass range were identified, which could act as species-specific biomarkers in complex microbial communities.

  9. The significance of gtf genes in caries expression: a rapid identification of Streptococcus mutans from dental plaque of child patients.

    Science.gov (United States)

    Mishra, Apurva; Pandey, Ramesh K; Manickam, Natesan

    2015-01-01

    Rapid phylogenetic and functional gene (gtfB) identification of S. mutans from the dental plaque derived from children. Dental plaque collected from fifteen patients of age group 7-12 underwent centrifugation followed by genomic DNA extraction for S. mutans. Genomic DNA was processed with S. mutans specific primers in suitable PCR condtions for phylogenetic and functional gene (gtfB) identification. The yield and results were confirmed by agarose gel electrophoresis. 1% agarose gel electrophoresis depicts the positive PCR amplification at 1,485 bp when compared with standard 1 kbp indicating the presence of S. mutans in the test sample. Another PCR reaction was set using gtfB primers specific for S. mutans for functional gene identification. 1.2% agarose gel electrophoresis was done and a positive amplication was observed at 192 bp when compared to 100 bp standards. With the advancement in molecular biology techniques, PCR based identification and quantification of the bacterial load can be done within hours using species-specific primers and DNA probes. Thus, this technique may reduce the laboratory time spend in conventional culture methods, reduces the possibility of colony identification errors and is more sensitive to culture techniques.

  10. Application of the LAMP Assay as a Diagnostic Technique for Rapid Identification of Thrips tabaci (Thysanoptera: Thripidae).

    Science.gov (United States)

    Fekrat, Lida; Zaki Aghl, Mohammad; Tahan, Vahid

    2015-06-01

    Rapid and accurate identification of potentially invasive taxa that may cause high economic losses or environmental damage is of critical importance. The onion thrips, Thrips tabaci Lindeman, ranks as one of the world's most destructive agricultural pests and commonly found in imported agricultural products and field samples, but is prone to undetected transport because of its minute size as well as cryptic behavior. Although traditional taxonomic methods are pretty useful in straightforward assignment of specimens to the genus Thrips, identification in the species level is much more difficult and requires expertise, knowledge, and experience. Furthermore, it is often difficult or impossible to identify or distinguish this species from other thrips by using material from other stages of development. Based on the foregoings, use of a molecular technique known as loop-mediated isothermal amplification (LAMP) as a rapid and robust alternative species diagnostic tool would be valuable. In this study, a relatively quick and simple method was used to detect the presence of onion thrips DNA rapidly and discriminate it from other species, by using material from different stages of development. Not only LAMP itself required less than 1 h to complete but also amounts of DNA as little as that recovered from a single specimen were adequate for the detection. Another advantage of this identification system is that nonspecialists will be able to make faster and cheaper identifications. © The Authors 2015. Published by Oxford University Press on behalf of Entomological Society of America. All rights reserved. For Permissions, please email: journals.permissions@oup.com.

  11. Rapid Species-level Identification of Salvias by Chemometric Processing of Ambient Ionisation Mass Spectrometry-derived Chemical Profiles.

    Science.gov (United States)

    Giffen, Justine E; Lesiak, Ashton D; Dane, A John; Cody, Robert B; Musah, Rabi A

    2017-01-01

    The Salvia genus contains numerous economically important plants that have horticultural, culinary and nutraceutical uses. They are often similar in appearance, making species determination difficult. Species identification of dried Salvia products is also challenging since distinguishing plant morphological features are no longer present. The development of a simple high-throughput method of analysis of fresh and dried Salvia leaves that would permit rapid species-level identification and detection of diagnostic biomarkers. Plant leaves were analysed in their native form by DART-MS without the need for any sample preparation steps. This furnished chemical fingerprints characteristic of each species. In the same experiment, in-source collision-induced dissociation was used to identify biomarkers. Biomarker presence was also independently confirmed by GC-MS. Chemometric processing of DART-MS profiles was performed by kernel discriminant analysis (KDA) and soft independent modelling of class analogy (SIMCA) to classify the fingerprints according to species. The approach was successful despite the occurrence of diurnal cycle and plant-age related chemical profile variations within species. In a single rapid experiment, the presence of essential oil biomarkers such as 3-carene, α-pinene, β-pinene, β-thujone, β-caryophyllene, camphor and borneol could be confirmed. The method was applied to rapid identification and differentiation of Salvia apiana, S. dominica, S. elegans, S. officinalis, S. farinacea and S. patens. Species-level identification of Salvia plant material could be accomplished by chemometric processing of DART-HRMS-derived chemical profiles of both fresh and dried Salvia material. Copyright © 2016 John Wiley & Sons, Ltd. Copyright © 2016 John Wiley & Sons, Ltd.

  12. Use of Fourier transform infrared spectroscopy (FTIR spectroscopy for rapid and accurate identification of Yeasts isolated from human and animals

    Directory of Open Access Journals (Sweden)

    M. Taha

    2013-06-01

    Full Text Available Rapid and accurate identification of yeast is increasingly important to stipulate the appropriate therapy thus reducing morbidity and mortality related to yeast infections. Vibrational spectroscopic techniques (infrared (IR and Raman could provide potential alternatives to conventional typing methods, because they constitute a rapid, inexpensive and highly specific spectroscopic fingerprint through-which microorganism can be identified. The present study evaluate (FTIR spectroscopy as a sensitive and effective assay for the identification of the most frequent yeast species isolated from human and animals. One hundred and twenty-eight yeasts isolated from infected human mouths/vaginas, chronic diseased cows, crop mycosis in chicken and soil contaminated with pigeon droppings were phenotypically identified. Using universal primers, ITS1/ITS4, we have amplified ITS1-5.8S-ITS2 rDNA regions for 39 yeast isolates as representative samples. The PCR products were digested with restriction enzyme MspI and examined by PCR-RFLP, which was an efficient technique for identification of Candida spp., Cryptococcus neoformans and Trichosporon asahii. Further, identification of the same 39 isolates were done by FTIR spectroscopy and considered as reference for other strains by comparison of their FTIR spectra. The current study has sharply demonstrated the significant spectral differences between the various examined species of Candida, Cryptococcus, Trichosporon, Rhodotorula and Geotrichum isolated from different sources. Decisively, our research has confirmed that FTIR spectroscopy is a promising diagnostic tool, because of its sensitivity, rapidity, high differentiation capacity and simplicity compared to conventional/molecular techniques.

  13. Exploring MALDI-TOF MS approach for a rapid identification of Mycobacterium avium ssp. paratuberculosis field isolates.

    Science.gov (United States)

    Ricchi, M; Mazzarelli, A; Piscini, A; Di Caro, A; Cannas, A; Leo, S; Russo, S; Arrigoni, N

    2017-03-01

    The aim of the study was to explore the suitability of matrix-assisted laser desorption/ionisation time-of-flight mass spectrometry (MALDI-TOF MS) for a rapid and correct identification of Mycobacterium avium ssp. paratuberculosis (MAP) field isolates. MALDI-TOF MS approach is becoming one of the most popular tests for the identification of intact bacterial cells which has been shown to be fast and reliable. For this purpose, 36 MAP field isolates were analysed through MALDI-TOF MS and the spectra compared with two different databases: one provided by the vendor of the system employed (Biotyper ver. 3·0; Bruker Daltonics) and a homemade database containing spectra from both tuberculous and nontuberculous Mycobacteria. Moreover, principal component analysis procedure was employed to confirm the ability of MALDI-TOF MS to discriminate between very closely related subspecies. Our results suggest MAP can be differentiated from other Mycobacterium species, both when the species are very close (M. intracellulare) and when belonging to different subspecies (M. avium ssp. avium and M. avium ssp. silvaticum). The procedure applied is fast, easy to perform, and achieves an earlier accurate species identification of MAP and nontuberculous Mycobacteria in comparison to other procedures. The gold standard test for the diagnosis of paratuberculosis is still isolation of MAP by cultural methods, but additional assays, such as qPCR and subculturing for determination of mycobactin dependency are required to confirm its identification. We have provided here evidence pertaining to the usefulness of MALDI-TOF MS approach for a rapid identification of this mycobacterium among other members of M. avium complex. © 2016 The Society for Applied Microbiology.

  14. Rapid Identification of Emerging Human-Pathogenic Sporothrix Species with Rolling Circle Amplification

    NARCIS (Netherlands)

    Rodrigues, Anderson M; Najafzadeh, Mohammad J; de Hoog, G Sybren; de Camargo, Zoilo P

    2015-01-01

    Sporothrix infections are emerging as an important human and animal threat among otherwise healthy patients, especially in Brazil and China. Correct identification of sporotrichosis agents is beneficial for epidemiological surveillance, enabling implementation of adequate public-health policies and

  15. Rapid identification of microorganisms from sterile body fluids by use of FilmArray.

    Science.gov (United States)

    Altun, Osman; Almuhayawi, Mohammed; Ullberg, Måns; Özenci, Volkan

    2015-02-01

    We evaluated the clinical performance of the FilmArray blood culture identification (BCID) panel in the identification of microorganisms from positive blood culture bottles inoculated with sterile body fluids. All organisms included in the FA BCID panel were accurately identified in 84/84 (100%) and 18/24 (75%) samples with mono- and polymicrobial growth, respectively. Copyright © 2015, American Society for Microbiology. All Rights Reserved.

  16. LV Barcoding: locality sensitive hashing-based tool for rapid species identification in DNA barcoding

    OpenAIRE

    Fan, Long; Chu, Ka Hou

    2014-01-01

    DNA barcoding has emerged as a cost-effective approach for species identification. However, the scarcity of tools used for searching the booming reference database becomes an obstacle, currently with BLAST as the only practical choice. Here, we propose a program - LV Barcoding - based on both the random hyperplane projection-based locality sensitive hashing method and the composition vector-based VIP Barcoding for fast species identification. The performance of LV Barcoding is assessed on the...

  17. Melting Temperature Mapping Method: A Novel Method for Rapid Identification of Unknown Pathogenic Microorganisms within Three Hours of Sample Collection.

    Science.gov (United States)

    Niimi, Hideki; Ueno, Tomohiro; Hayashi, Shirou; Abe, Akihito; Tsurue, Takahiro; Mori, Masashi; Tabata, Homare; Minami, Hiroshi; Goto, Michihiko; Akiyama, Makoto; Yamamoto, Yoshihiro; Saito, Shigeru; Kitajima, Isao

    2015-07-28

    Acquiring the earliest possible identification of pathogenic microorganisms is critical for selecting the appropriate antimicrobial therapy in infected patients. We herein report the novel "melting temperature (Tm) mapping method" for rapidly identifying the dominant bacteria in a clinical sample from sterile sites. Employing only seven primer sets, more than 100 bacterial species can be identified. In particular, using the Difference Value, it is possible to identify samples suitable for Tm mapping identification. Moreover, this method can be used to rapidly diagnose the absence of bacteria in clinical samples. We tested the Tm mapping method using 200 whole blood samples obtained from patients with suspected sepsis, 85% (171/200) of which matched the culture results based on the detection level. A total of 130 samples were negative according to the Tm mapping method, 98% (128/130) of which were also negative based on the culture method. Meanwhile, 70 samples were positive according to the Tm mapping method, and of the 59 suitable for identification, 100% (59/59) exhibited a "match" or "broad match" with the culture or sequencing results. These findings were obtained within three hours of whole blood collection. The Tm mapping method is therefore useful for identifying infectious diseases requiring prompt treatment.

  18. Lncident: A Tool for Rapid Identification of Long Noncoding RNAs Utilizing Sequence Intrinsic Composition and Open Reading Frame Information

    Directory of Open Access Journals (Sweden)

    Siyu Han

    2016-01-01

    Full Text Available More and more studies have demonstrated that long noncoding RNAs (lncRNAs play critical roles in diversity of biological process and are also associated with various types of disease. How to rapidly identify lncRNAs and messenger RNA is the fundamental step to uncover the function of lncRNAs identification. Here, we present a novel method for rapid identification of lncRNAs utilizing sequence intrinsic composition features and open reading frame information based on support vector machine model, named as Lncident (LncRNAs identification. The 10-fold cross-validation and ROC curve are used to evaluate the performance of Lncident. The main advantage of Lncident is high speed without the loss of accuracy. Compared with the exiting popular tools, Lncident outperforms Coding-Potential Calculator, Coding-Potential Assessment Tool, Coding-Noncoding Index, and PLEK. Lncident is also much faster than Coding-Potential Calculator and Coding-Noncoding Index. Lncident presents an outstanding performance on microorganism, which offers a great application prospect to the analysis of microorganism. In addition, Lncident can be trained by users’ own collected data. Furthermore, R package and web server are simultaneously developed in order to maximize the convenience for the users. The R package “Lncident” can be easily installed on multiple operating system platforms, as long as R is supported.

  19. Changes in Sensory Evoked Responses Coincide with Rapid Improvement in Speech Identification Performance

    Science.gov (United States)

    Alain, Claude; Campeanu, Sandra; Tremblay, Kelly

    2010-01-01

    Perceptual learning is sometimes characterized by rapid improvements in performance within the first hour of training (fast perceptual learning), which may be accompanied by changes in sensory and/or response pathways. Here, we report rapid physiological changes in the human auditory system that coincide with learning during a 1-hour test session…

  20. Rapid detection and identification of Stachybotrys and Chaetomium species using tissue PCR analysis

    DEFF Research Database (Denmark)

    Lewinska, Anna Malgorzata; Peuhkuri, Ruut Hannele; Rode, Carsten

    2016-01-01

    Indoor fungi are a worldwide problem causing negative health effects for infected building's occupants and even deterioration of building structures. Different fungal species affect buildings and their inhabitants differently. Therefore, rapid and accurate identification of fungi to the species l...

  1. Tip-tilt disturbance model identification for Kalman-based control scheme: application to XAO and ELT systems.

    Science.gov (United States)

    Meimon, Serge; Petit, Cyril; Fusco, Thierry; Kulcsar, Caroline

    2010-11-01

    Adaptive optics (AO) systems have to correct tip-tilt (TT) disturbances down to a fraction of the diffraction-limited spot. This becomes a key issue for very or extremely large telescopes affected by mechanical vibration peaks or wind shake effects. Linear quadratic Gaussian (LQG) control achieves optimal TT correction when provided with the temporal model of the disturbance. We propose a nonsupervised identification procedure that does not require any auxiliary system or loop opening and validate it on synthetic profile as well as on experimental data.

  2. Rapid detection and identification of viral and bacterial fish pathogens using a DNA array‐based multiplex assay

    DEFF Research Database (Denmark)

    Lievens, B.; Frans, I.; Heusdens, C.

    2011-01-01

    for the simultaneous detection and identification of all cyprinid herpesviruses (CyHV‐1, CyHV‐2 and CyHV‐3) and some of the most important fish pathogenic Flavobacterium species, including F. branchiophilum, F. columnare and F. psychrophilum. For virus identification, the DNA polymerase and helicase genes were......Fish diseases can be caused by a variety of diverse organisms, including bacteria, fungi, viruses and protozoa, and pose a universal threat to the ornamental fish industry and aquaculture. The lack of rapid, accurate and reliable means by which fish pathogens can be detected and identified has been...... one of the main limitations in fish pathogen diagnosis and fish disease management and has consequently stimulated the search for alternative diagnostic techniques. Here, we describe a method based on multiplex and broad‐range PCR amplification combined with DNA array hybridization...

  3. A Rapid and Simple LC-MS Method Using Collagen Marker Peptides for Identification of the Animal Source of Leather.

    Science.gov (United States)

    Kumazawa, Yuki; Taga, Yuki; Iwai, Kenji; Koyama, Yoh-Ichi

    2016-08-03

    Identification of the animal source of leather is difficult using traditional methods, including microscopic observation and PCR. In the present study, a LC-MS method was developed for detecting interspecies differences in the amino acid sequence of type I collagen, which is a major component of leather, among six animals (cattle, horse, pig, sheep, goat, and deer). After a dechroming procedure and trypsin digestion, six tryptic peptides of type I collagen were monitored by LC-MS in multiple reaction monitoring mode for the animal source identification using the patterns of the presence or absence of the marker peptides. We analyzed commercial leathers from various production areas using this method, and found some leathers in which the commercial label disagreed with the identified animal source. Our method enabled rapid and simple leather certification and could be applied to other animals whether or not their collagen sequences are available in public databases.

  4. Rapid and Accurate Identification of Animal Species in Natural Leather Goods by Liquid Chromatography/Mass Spectrometry.

    Science.gov (United States)

    Izuchi, Yukari; Takashima, Tsuneo; Hatano, Naoya

    2016-01-01

    The demand for leather goods has grown globally in recent years. Industry revenue is forecast to reach $91.2 billion by 2018. There is an ongoing labelling problem in the leather items market, in that it is currently impossible to identify the species that a given piece of leather is derived from. To address this issue, we developed a rapid and simple method for the specific identification of leather derived from cattle, horses, pigs, sheep, goats, and deer by analysing peptides produced by the trypsin-digestion of proteins contained in leather goods using liquid chromatography/mass spectrometry. We determined species-specific amino acid sequences by liquid chromatography/tandem mass spectrometry analysis using the Mascot software program and demonstrated that collagen α-1(I), collagen α-2(I), and collagen α-1(III) from the dermal layer of the skin are particularly useful in species identification.

  5. Rapid Sanger sequencing of the 16S rRNA gene for identification of some common pathogens.

    Directory of Open Access Journals (Sweden)

    Linxiang Chen

    Full Text Available Conventional Sanger sequencing remains time-consuming and laborious. In this study, we developed a rapid improved sequencing protocol of 16S rRNA for pathogens identification by using a new combination of SYBR Green I real-time PCR and Sanger sequencing with FTA® cards. To compare the sequencing quality of this method with conventional Sanger sequencing, 12 strains, including three kinds of strains (1 reference strain and 3 clinical strains, which were previously identified by biochemical tests, which have 4 Pseudomonas aeruginosa, 4 Staphyloccocus aureus and 4 Escherichia coli, were targeted. Additionally, to validate the sequencing results and bacteria identification, expanded specimens with 90 clinical strains, also comprised of the three kinds of strains which included 30 samples respectively, were performed as just described. The results showed that although statistical differences (P<0.05 were found in sequencing quality between the two methods, their identification results were all correct and consistent. The workload, the time consumption and the cost per batch were respectively light versus heavy, 8 h versus 11 h and $420 versus $400. In the 90 clinical strains, all of the Pseudomonas aeruginosa and Staphyloccocus aureus strains were correctly identified, but only 26.7% of the Escherichia coli strains were recognized as Escherichia coli, while 33.3% as Shigella sonnei and 40% as Shigella dysenteriae. The protocol described here is a rapid, reliable, stable and convenient method for 16S rRNA sequencing, and can be used for Pseudomonas aeruginosa and Staphyloccocus aureus identification, yet it is not completely suitable for discriminating Escherichia coli and Shigella strains.

  6. Rapid identification of genetic modifications in Bacillus anthracis using whole genome draft sequences generated by 454 pyrosequencing.

    Directory of Open Access Journals (Sweden)

    Peter E Chen

    Full Text Available BACKGROUND: The anthrax letter attacks of 2001 highlighted the need for rapid identification of biothreat agents not only for epidemiological surveillance of the intentional outbreak but also for implementing appropriate countermeasures, such as antibiotic treatment, in a timely manner to prevent further casualties. It is clear from the 2001 cases that survival may be markedly improved by administration of antimicrobial therapy during the early symptomatic phase of the illness; i.e., within 3 days of appearance of symptoms. Microbiological detection methods are feasible only for organisms that can be cultured in vitro and cannot detect all genetic modifications with the exception of antibiotic resistance. Currently available immuno or nucleic acid-based rapid detection assays utilize known, organism-specific proteins or genomic DNA signatures respectively. Hence, these assays lack the ability to detect novel natural variations or intentional genetic modifications that circumvent the targets of the detection assays or in the case of a biological attack using an antibiotic resistant or virulence enhanced Bacillus anthracis, to advise on therapeutic treatments. METHODOLOGY/PRINCIPAL FINDINGS: We show here that the Roche 454-based pyrosequencing can generate whole genome draft sequences of deep and broad enough coverage of a bacterial genome in less than 24 hours. Furthermore, using the unfinished draft sequences, we demonstrate that unbiased identification of known as well as heretofore-unreported genetic modifications that include indels and single nucleotide polymorphisms conferring antibiotic and phage resistances is feasible within the next 12 hours. CONCLUSIONS/SIGNIFICANCE: Second generation sequencing technologies have paved the way for sequence-based rapid identification of both known and previously undocumented genetic modifications in cultured, conventional and newly emerging biothreat agents. Our findings have significant implications in

  7. Rapid and accurate identification of isolates of Candida species by melting peak and melting curve analysis of the internally transcribed spacer region 2 fragment (ITS2-MCA)

    NARCIS (Netherlands)

    Decat, E.; van Mechelen, E.; Saerens, B.; Vermeulen, S.J.T.; Boekhout, T.; de Blaiser, S.; Vaneechoutte, M.; Deschaght, P.

    2013-01-01

    Rapid identification of clinically important yeasts can facilitate the initiation of anti-fungal therapy, since susceptibility is largely species-dependent. We evaluated melting peak and melting curve analysis of the internally transcribed spacer region 2 fragment (ITS2-MCA) as an identification

  8. Rapid identification of bacterial biofilms and biofilm wound models using a multichannel nanosensor.

    Science.gov (United States)

    Li, Xiaoning; Kong, Hao; Mout, Rubul; Saha, Krishnendu; Moyano, Daniel F; Robinson, Sandra M; Rana, Subinoy; Zhang, Xinrong; Riley, Margaret A; Rotello, Vincent M

    2014-12-23

    Identification of infectious bacteria responsible for biofilm-associated infections is challenging due to the complex and heterogeneous biofilm matrix. To address this issue and minimize the impact of heterogeneity on biofilm identification, we developed a gold nanoparticle (AuNP)-based multichannel sensor to detect and identify biofilms based on their physicochemical properties. Our results showed that the sensor can discriminate six bacterial biofilms including two composed of uropathogenic bacteria. The capability of the sensor was further demonstrated through discrimination of biofilms in a mixed bacteria/mammalian cell in vitro wound model.

  9. Increased Sensitivity of a New Coagglutination Test for Rapid Identification of Haemophilus influenzae Type b

    OpenAIRE

    Grasso, Robert J.; West, Loyd A.; Holbrook, Nikki J.; Halkias, Demetrios G.; Paradise, Lois J.; Friedman, Herman

    1981-01-01

    A newly developed rapid coagglutination test for identifying Haemophilus influenzae type b organisms isolated from clinical specimens correlated 100% with the slide agglutination test but was 100- to 200-fold more sensitive.

  10. Liposome immunoassay for rapid identification of group A streptococci directly from throat swabs.

    OpenAIRE

    Gerber, M. A.; Randolph, M. F.; DeMeo, K K

    1990-01-01

    The Q Test Strep (Becton Dickinson and Co., Franklin Lakes, N.J.) is a new solid-phase liposome immunoassay for the rapid diagnosis of group A beta-hemolytic streptococcal pharyngitis. Compared with blood agar plate cultures, the Q Test Strep had a sensitivity of 91%, a specificity of 83%, a positive predictive value of 88%, and a negative predictive value of 87%. Liposome technology can be used to facilitate the rapid diagnosis of group A beta-hemolytic streptococcal pharyngitis.

  11. Rapid and correct identification of intestinal Bacteroides spp. with chromosomal DNA probes by whole-cell dot blot hybridization

    Energy Technology Data Exchange (ETDEWEB)

    Morotomi, M.; Ohno, T.; Mutai, M.

    1988-05-01

    A dot blot hybridization procedure with /sup 32/P-labeled whole chromosomal DNA of the type strains as probes was developed as a rapid and simple method for identification of intestinal Bacteroides species. Bacterial cells were fixed onto membrane filters by slight suction, treated with 0.5 N NaOH, and hybridized with these probes. Of 65 Bacteroides strains isolated from 19 human fecal specimens, which were identified as B. fragilis, B. thetaiotaomicron, B. ovatus, B. caccae, B. uniformis, B. stercoris, B. vulgatus, B. distasonis, and B. merdae by conventional phenotypic characterization, 62 (95%) were correctly identified with this hybridization procedure.

  12. Identification and characterization of hierarchical structures in dog breeding schemes, a novel method applied to the Norfolk Terrier.

    Science.gov (United States)

    Wellmann, R; Bennewitz, J

    2011-12-01

    Popular sires, a limited population size, and the founder event are widely considered the main reasons for the low genetic diversity observed in many dog breeds. However, these factors have had only a small role in the historic decrease in diversity observed in the Norfolk Terrier breed. We show that the decrease in this breed has been mainly due to large, popular kennels. Dogs from these kennels have, on average, larger genetic contributions to subsequent generations than others. A test for the presence of a popular kennel effect is proposed and applied (P dog breeding schemes, with an asymmetric gene flow predominantly from the nuclei toward the main population. Possible reasons for this structure and implications for future population management are discussed. The main reason is probably that the breed type was established by large, popular kennels and that small kennel breeders used their stud dogs to benefit from the achievements of the popular kennels. Many kennels, however, were unable to make their own substantial genetic contributions to the breed.

  13. Rapid identification and simultaneous analysis of multiple constituents from Rheum tanguticum Maxim. ex Balf. by UPLC/Q-TOF-MS.

    Science.gov (United States)

    Gao, Liang-Liang; Guo, Tao; Xu, Xu-Dong; Yang, Jun-Shan

    2017-07-01

    Rhubarb contains biologically active compounds such as anthraquinones, anthrones, stilbenes and tannins. A rapid and efficient UPLC/Q-TOF-MS/MS method was developed and applied towards identifying the constituents of Rheum tanguticum Maxim. ex Balf. for the first time. Chemical constituents were separated and investigated by UPLC/Q-TOF-MS/MS in the negative ion mode. The ESI-MS(2) fragmentation pathways of four types of compounds were interpreted, providing a very useful guidance for the characterisation of different types of compounds. Based on the exact mass information, fragmentation characteristic and LC retention time of 7 reference standards, 30 constituents were tentatively identified from the methanol extract of R. tanguticum. Among them, seven compounds were described for the first time from R. tanguticum and two from the genus Rheum were described for the first time. The analytical tool used here is valuable for the rapid separation and identification of multiple and minor constituents in methanol extracts of R. tanguticum.

  14. Poly(4-vinylphenol) gate insulator with cross-linking using a rapid low-power microwave induction heating scheme for organic thin-film-transistors

    OpenAIRE

    Ching-Lin Fan; Ming-Chi Shang; Mao-Yuan Hsia; Shea-Jue Wang; Bohr-Ran Huang; Win-Der Lee

    2016-01-01

    A Microwave-Induction Heating (MIH) scheme is proposed for the poly(4-vinylphenol) (PVP) gate insulator cross-linking process to replace the traditional oven heating cross-linking process. The cross-linking time is significantly decreased from 1 h to 5 min by heating the metal below the PVP layer using microwave irradiation. The necessary microwave power was substantially reduced to about 50 W by decreasing the chamber pressure. The MIH scheme is a good candidate to replace traditional therma...

  15. Rapid identification of allergenic and pathogenic molds in environmental air by an oligonucleotide array.

    Science.gov (United States)

    Hung, Wen-Tsung; Su, Shu-Li; Shiu, Lin-Yi; Chang, Tsung C

    2011-04-13

    Airborne fungi play an important role in causing allergy and infections in susceptible people. Identification of these fungi, based on morphological characteristics, is time-consuming, expertise-demanding, and could be inaccurate. We developed an oligonucleotide array that could accurately identify 21 important airborne fungi (13 genera) that may cause adverse health problems. The method consisted of PCR amplification of the internal transcribed spacer (ITS) regions, hybridization of the PCR products to a panel of oligonucleotide probes immobilized on a nylon membrane, and detection of the hybridization signals with alkaline phosphatase-conjugated antibodies. A collection of 72 target and 66 nontarget reference strains were analyzed by the array. Both the sensitivity and specificity of the array were 100%, and the detection limit was 10 pg of genomic DNA per assay. Furthermore, 70 fungal isolates recovered from air samples were identified by the array and the identification results were confirmed by sequencing of the ITS and D1/D2 domain of the large-subunit RNA gene. The sensitivity and specificity of the array for identification of the air isolates was 100% (26/26) and 97.7% (43/44), respectively. Identification of airborne fungi by the array was cheap and accurate. The current array may contribute to decipher the relationship between airborne fungi and adverse health effect.

  16. A PCR-based strategy for simple and rapid identification of rough presumptive Salmonella isolates

    DEFF Research Database (Denmark)

    Hoorfar, Jeffrey; Baggesen, Dorte Lau; Porting, P.H.

    1999-01-01

    The purpose of the present study was to investigate the application of ready-to-go Salmonella PCR tests, based on dry chemistry, for final identification of rough presumptive Salmonella isolates. The results were compared with two different biotyping methods performed at two different laboratories...

  17. Rapid identification of allergenic and pathogenic molds in environmental air by an oligonucleotide array

    Directory of Open Access Journals (Sweden)

    Shiu Lin-Yi

    2011-04-01

    Full Text Available Abstract Background Airborne fungi play an important role in causing allergy and infections in susceptible people. Identification of these fungi, based on morphological characteristics, is time-consuming, expertise-demanding, and could be inaccurate. Methods We developed an oligonucleotide array that could accurately identify 21 important airborne fungi (13 genera that may cause adverse health problems. The method consisted of PCR amplification of the internal transcribed spacer (ITS regions, hybridization of the PCR products to a panel of oligonucleotide probes immobilized on a nylon membrane, and detection of the hybridization signals with alkaline phosphatase-conjugated antibodies. Results A collection of 72 target and 66 nontarget reference strains were analyzed by the array. Both the sensitivity and specificity of the array were 100%, and the detection limit was 10 pg of genomic DNA per assay. Furthermore, 70 fungal isolates recovered from air samples were identified by the array and the identification results were confirmed by sequencing of the ITS and D1/D2 domain of the large-subunit RNA gene. The sensitivity and specificity of the array for identification of the air isolates was 100% (26/26 and 97.7% (43/44, respectively. Conclusions Identification of airborne fungi by the array was cheap and accurate. The current array may contribute to decipher the relationship between airborne fungi and adverse health effect.

  18. Rapid identification of bacteria in blood cultures by using fluorescently labeled oligonucleotide probes

    NARCIS (Netherlands)

    Jansen, GJ; Mooibroek, M; Idema, J; Harmsen, HJM; Welling, GW; Degener, JE

    The applicability of whole-cell hybridization for the identification of pathogenic bacteria in blood from septic patients was examined. Oligonucleotide probes, fluorescently labeled with fluorescein isothiocyanate, directed against the variable regions of the 16S rRNAs of the following bacterial

  19. Development of Conductive Polymer Analysis for the Rapid Detection and Identification of Phytopathogenic Microbes

    Science.gov (United States)

    A. Dan Wilson; D.G. Lester; C.S. Oberle

    2004-01-01

    Conductive polymer analysis, a type of electronic aroma detection technology, was evaluated for its efficacy in the detection, identification, and discrimination of plant-pathogenic microorganisms on standardized media and in diseased plant tissues. The method is based on the acquisition of a diagnostic electronic fingerprint derived from multisensor responses to...

  20. Rapid and inexpensive body fluid identification by RNA profiling-based multiplex High Resolution Melt (HRM) analysis.

    Science.gov (United States)

    Hanson, Erin K; Ballantyne, Jack

    2013-01-01

    Positive identification of the nature of biological material present on evidentiary items can be crucial for understanding the circumstances surrounding a crime. However, traditional protein-based methods do not permit the identification of all body fluids and tissues, and thus molecular based strategies for the conclusive identification of all forensically relevant biological fluids and tissues need to be developed. Messenger RNA (mRNA) profiling is an example of such a molecular-based approach. Current mRNA body fluid identification assays involve capillary electrophoresis (CE) or quantitative RT-PCR (qRT-PCR) platforms, each with its own limitations. Both platforms require the use of expensive fluorescently labeled primers or probes. CE-based assays require separate amplification and detection steps thus increasing the analysis time. For qRT-PCR assays, only 3-4 markers can be included in a single reaction since each requires a different fluorescent dye. To simplify mRNA profiling assays, and reduce the time and cost of analysis, we have developed single- and multiplex body fluid High Resolution Melt (HRM) assays for the identification of common forensically relevant biological fluids and tissues. The incorporated biomarkers include IL19 (vaginal secretions), IL1F7 (skin), ALAS2 (blood), MMP10 (menstrual blood), HTN3 (saliva) and TGM4 (semen).  The HRM assays require only unlabeled PCR primers and a single saturating intercalating fluorescent dye (Eva Green). Each body-fluid-specific marker can easily be identified by the presence of a distinct melt peak. Usually, HRM assays are used to detect variants or isoforms for a single gene target. However, we have uniquely developed duplex and triplex HRM assays to permit the simultaneous detection of multiple targets per reaction. Here we describe the development and initial performance evaluation of the developed HRM assays. The results demonstrate the potential use of HRM assays for rapid, and relatively inexpensive

  1. Matrix-assisted laser desorption/ionization time-of-flight mass spectrometry for rapid identification of fungal rhinosinusitis pathogens.

    Science.gov (United States)

    Huang, Yanfei; Wang, Jinglin; Zhang, Mingxin; Zhu, Min; Wang, Mei; Sun, Yufeng; Gu, Haitong; Cao, Jingjing; Li, Xue; Zhang, Shaoya; Lu, Xinxin

    2017-03-01

    Filamentous fungi are among the most important pathogens, causing fungal rhinosinusitis (FRS). Current laboratory diagnosis of FRS pathogens mainly relies on phenotypic identification by culture and microscopic examination, which is time consuming and expertise dependent. Although matrix-assisted laser desorption/ionization time-of-flight (MALDI-TOF) MS has been employed to identify various fungi, its efficacy in the identification of FRS fungi is less clear. A total of 153 FRS isolates obtained from patients were analysed at the Clinical Laboratory at the Beijing Tongren Hospital affiliated to the Capital Medical University, between January 2014 and December 2015. They were identified by traditional phenotypic methods and Bruker MALDI-TOF MS (Bruker, Biotyper version 3.1), respectively. Discrepancies between the two methods were further validated by sequencing. Among the 153 isolates, 151 had correct species identification using MALDI-TOF MS (Bruker, Biot 3.1, score ≥2.0 or 2.3). MALDI-TOF MS enabled identification of some very closely related species that were indistinguishable by conventional phenotypic methods, including 1/10 Aspergillus versicolor, 3/20 Aspergillus flavus, 2/30 Aspergillus fumigatus and 1/20 Aspergillus terreus, which were misidentified by conventional phenotypic methods as Aspergillus nidulans, Aspergillus oryzae, Aspergillus japonicus and Aspergillus nidulans, respectively. In addition, 2/2 Rhizopus oryzae and 1/1 Rhizopus stolonifer that were identified only to the genus level by the phenotypic method were correctly identified by MALDI-TOF MS. MALDI-TOF MS is a rapid and accurate technique, and could replace the conventional phenotypic method for routine identification of FRS fungi in clinical microbiology laboratories.

  2. Rapid identification of bacillus anthracis spores in suspicious powder samples by using matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS)

    NARCIS (Netherlands)

    Dybwad, M.; Laaken, A.L. van der; Blatny, J.M.; Paauw, A.

    2013-01-01

    Rapid and reliable identification of Bacillus anthracis spores in suspicious powders is important to mitigate the safety risks and economic burdens associated with such incidents. The aim of this study was to develop and validate a rapid and reliable laboratory- based matrix-assisted laser

  3. Rapid identification of HBB gene mutations by high-resolution melting analysis.

    Science.gov (United States)

    Shih, Hung-Chang; Er, Tze-Kiong; Chang, Tien-Jye; Chang, Ya-Sian; Liu, Ta-Chih; Chang, Jan-Gowth

    2009-11-01

    This study was undertaken to identify HBB gene mutation. Herein we evaluated high-resolution melting analysis in the identification of HBB mutations. We have successfully established a diagnostic strategy for identifying HBB gene mutations including c.-78A>G, c.-79A>G, c.2T>G, c.79_80insT, c.84_85insC, c.123_124insT, c.125_128delTCTT, c.130 G>T, c.170G>A, c.216_217ins A and c.316-197 C>T from wild-type DNA using HRM analysis. The results of HRM analysis were confirmed by direct DNA sequencing. In summary, we report that HRM analysis is an appealing technique for the identification of HBB mutations. We also believe that HRM can be used as a method for prenatal diagnosis of beta-thalassemia.

  4. Rapid identification of pathogens from positive blood cultures by multiplex polymerase chain reaction using the FilmArray system.

    Science.gov (United States)

    Blaschke, Anne J; Heyrend, Caroline; Byington, Carrie L; Fisher, Mark A; Barker, Elizabeth; Garrone, Nicholas F; Thatcher, Stephanie A; Pavia, Andrew T; Barney, Trenda; Alger, Garrison D; Daly, Judy A; Ririe, Kirk M; Ota, Irene; Poritz, Mark A

    2012-12-01

    Sepsis is a leading cause of death. Rapid and accurate identification of pathogens and antimicrobial resistance directly from blood culture could improve patient outcomes. The FilmArray® (FA; Idaho Technology, Salt Lake City, UT, USA) Blood Culture (BC) panel can identify >25 pathogens and 4 antibiotic resistance genes from positive blood cultures in 1 h. We compared a development version of the panel to conventional culture and susceptibility testing on 102 archived blood cultures from adults and children with bacteremia. Of 109 pathogens identified by culture, 95% were identified by FA. Among 111 prospectively collected blood cultures, the FA identified 84 (91%) of 92 pathogens covered by the panel. Among 25 Staphylococcus aureus and 21 Enterococcus species detected, FA identified all culture-proven methicillin-resistant S. aureus and vancomycin-resistant enterococci. The FA BC panel is an accurate method for the rapid identification of pathogens and resistance genes from blood culture. Copyright © 2012 Elsevier Inc. All rights reserved.

  5. Rapid sex identification method of spinach (Spinacia oleracea L.) in the vegetative stage using loop-mediated isothermal amplification.

    Science.gov (United States)

    Fujita, Naoko; Ayukawa, Yu; Fuke, Mitsutoshi; Teraoka, Tohru; Watanabe, Kyoko; Arie, Tsutomu; Komatsu, Ken

    2017-01-01

    A LAMP-mediated, simple and rapid method for sex identification in spinach was developed. Nutrient compositional analysis showed a higher iron content in male than female plants. Spinach (Spinacia oleracea L.) is a dioecious plant with its sex determined by the XY system. Male and female floral organs differ morphologically, but plants do not differ in the vegetative stage before flowering. PCR with Y chromosome markers has been used to determine the sex of dioecious plants before flowering. In this study, we developed a genotype-specific loop-mediated isothermal amplification (LAMP) for sex identification of individual vegetative-stage spinach plants, using primers designed for the genomic region flanked by male-specific markers. LAMP could specifically detect spinach males. The method was further modified to omit DNA purification and use just an aliquot of crude leaf extract homogenized in water. We compared the nutrient composition of males and females, finding higher amounts of iron in the males. Our method could therefore be used for rapidly discriminating male plants in the field, which is useful for efficient hybrid breeding.

  6. Rapid identification and quantitative analysis of chemical constituents of Gentiana veitchiorum by UHPLC-PDA-QTOF-MS

    Directory of Open Access Journals (Sweden)

    Shan Li

    Full Text Available ABSTRACT Gentiana veitchiorum Hemsl., Gentianaceae, a traditional Tibetan medicine, was used for the treatment of liver jaundice with damp-heat pathogen, as well as for headache and chronic pharyngitis. A rapid ultra-performance liquid chromatography, photodiode array detector, quadrupole time-of-flight mass spectrometry method was developed for the fast and accurate identification and quantification of the chemical constituents of G. veitchiorum. In fact, eighteen compounds were detected and identified on the basis of their mass spectra, fragment characteristics and comparison with published data. Especially, the MS fragmentation pathways of iridoid glycosides and flavone C-glycosides were illustrated. Five compounds among them were quantified by UHPLC-PDA, including swertiamarin, gentiopicroside, sweroside, isoorientin, and isovitexin. The proposed method was then validated based on the analyses of linearity, accuracy, precision, and recovery. The overall recoveries for the five analytes ranged from 96.54% to 100.81%, with RSD from 1.05% to 1.82%. In addition, ten batches of G. veitchiorum from different areas were also analyzed. The developed method was rapid and reliable for both identification and quantification of the chemical constituents of G. veitchiorum, especially for simultaneous qualitative and quantitative analysis of iridoid glycosides and flavone C-glycosides.

  7. Development of loop-mediated isothermal amplification (LAMP assay for rapid and sensitive identification of ostrich meat.

    Directory of Open Access Journals (Sweden)

    Amir Abdulmawjood

    Full Text Available Animal species identification is one of the primary duties of official food control. Since ostrich meat is difficult to be differentiated macroscopically from beef, therefore new analytical methods are needed. To enforce labeling regulations for the authentication of ostrich meat, it might be of importance to develop and evaluate a rapid and reliable assay. In the present study, a loop-mediated isothermal amplification (LAMP assay based on the cytochrome b gene of the mitochondrial DNA of the species Struthio camelus was developed. The LAMP assay was used in combination with a real-time fluorometer. The developed system allowed the detection of 0.01% ostrich meat products. In parallel, a direct swab method without nucleic acid extraction using the HYPLEX LPTV buffer was also evaluated. This rapid processing method allowed detection of ostrich meat without major incubation steps. In summary, the LAMP assay had excellent sensitivity and specificity for detecting ostrich meat and could provide a sampling-to-result identification-time of 15 to 20 minutes.

  8. Comparative analysis of simulated candidemia using two different blood culture systems and the rapid identification of Candida albicans.

    Science.gov (United States)

    Park, Bo Rae G; Kim, Tae-Hyoung; Kim, Hye Ryoun; Lee, Mi-Kyung

    2011-01-01

    The goal of this study was to determine the time to detection of Candida species isolates using the two most commonly used automated blood culture systems, and to evaluate rapid, widely available methods for the presumptive identification of C. albicans. Candidemia models of eight commonly detected Candida species were prepared using ATCC standards. The times to detection were evaluated using the BACTEC 9240 (Becton Dickinson) and BacT/Alert 3D (bioMerieux) automated blood culture systems. The presence of pseudohyphae clusters was examined by Gram staining and wet preparation. Germ tube tests were performed directly from blood culture bottles. All samples were cultured on blood agar plates and macroscopically examined for the presence of an irregular margin (spiking). Most Candida species (6/8) except C. glabrata and C. krusei grew more rapidly in aerobic than in anaerobic conditions. Clusters of pseudohyphae were observed in cultures of C. albicans and C. tropicalis. All culture bottles positive for C. albicans were positive by the germ tube test and macroscopically showed 'spiking.' Aerobic and anaerobic blood culture systems can effectively detect candidemia. Furthermore, the direct germ tube test may be the most useful available morphological presumptive identification method for C. albicans.

  9. Implementation of an FTIR spectral library of 486 filamentous fungi strains for rapid identification of molds.

    Science.gov (United States)

    Lecellier, A; Gaydou, V; Mounier, J; Hermet, A; Castrec, L; Barbier, G; Ablain, W; Manfait, M; Toubas, D; Sockalingum, G D

    2015-02-01

    Filamentous fungi may cause food and feed spoilage and produce harmful metabolites to human and animal health such as mycotoxins. Identification of fungi using conventional phenotypic methods is time-consuming and molecular methods are still quite expensive and require specific laboratory skills. In the last two decades, it has been shown that Fourier transform infrared (FTIR) spectroscopy was an efficient tool for microorganism identification. The aims of this study were to use a simple protocol for the identification of filamentous fungi using FTIR spectroscopy coupled with a partial least squares discriminant analysis (PLS-DA), to implement a procedure to validate the obtained results, and to assess the transferability of the method and database. FTIR spectra of 486 strains (43 genera and 140 species) were recorded. An IR spectral database built with 288 strains was used to identify 105 different strains. It was found that 99.17% and 92.3% of spectra derived from these strains were correctly assigned at the genus and species levels, respectively. The establishment of a score and a threshold permitted to validate 80.79% of the results obtained. A standardization function (SF) was also implemented and tested on FTIR data from another instrument on a different site and permitted to increase the percentage of well predicted spectra for this set from 72.15% to 89.13%. This study confirms the good performance of high throughput FTIR spectroscopy for fungal identification using a spectral library of molds of industrial relevance. Copyright © 2014 Elsevier Ltd. All rights reserved.

  10. Surface Enhanced Raman Spectroscopy for the Rapid Detection and Identification of Microbial Pathogens in Human Serum

    Science.gov (United States)

    2014-12-11

    those of the authors and do not necessarily reflect the official policy or position of the Department of the Navy, Department of Defense, nor the U.S...the medical field, Raman spectroscopy is under investigation for use in prediction of preterm birth [10], identification of basal cell carcinoma [11...in peak intensity at 735 cm -1 was considered positive for microbial presence. Variability in Raman signal across SERS spectra was determined by

  11. A multiplex PCR for rapid identification of Brassica species in the triangle of U

    OpenAIRE

    Koh, Joshua C. O.; Denise M Barbulescu; Norton, Sally; Redden, Bob; Salisbury, Phil A.; Kaur, Sukhjiwan; Cogan, Noel; Slater, Anthony T.

    2017-01-01

    Background Within the Brassicaceae, six species from the genus Brassica are widely cultivated throughout the world as oilseed, condiment, fodder or vegetable crops. The genetic relationships among the six Brassica species are described by U?s triangle model. Extensive shared traits and diverse morphotypes among Brassica species make identification and classification based on phenotypic data alone challenging and unreliable, especially when dealing with large germplasm collections. Consequentl...

  12. Rapid identification of Candida albicans by using Albicans ID and fluoroplate agar plates.

    OpenAIRE

    Rousselle, P.; Freydiere, A. M.; Couillerot, P J; De Montclos, H.; Gille, Y.

    1994-01-01

    Two commercially available agar media, Albicans ID and Fluoroplate, that use a chromogenic or a fluorogenic substrate for the detection and identification of Candida albicans were evaluated. From 1,006 clinical samples containing 723 yeast strains, 352 C. albicans strains were detected with either of the two media. The sensitivity of each of the two media was 93.8% and the specificity was 98.6%, with five false-positive reactions for Candida tropicalis and no false-negative reactions.

  13. Rapid identification of causal mutations in tomato EMS populations via mapping-by-sequencing.

    Science.gov (United States)

    Garcia, Virginie; Bres, Cécile; Just, Daniel; Fernandez, Lucie; Tai, Fabienne Wong Jun; Mauxion, Jean-Philippe; Le Paslier, Marie-Christine; Bérard, Aurélie; Brunel, Dominique; Aoki, Koh; Alseekh, Saleh; Fernie, Alisdair R; Fraser, Paul D; Rothan, Christophe

    2016-12-01

    The tomato is the model species of choice for fleshy fruit development and for the Solanaceae family. Ethyl methanesulfonate (EMS) mutants of tomato have already proven their utility for analysis of gene function in plants, leading to improved breeding stocks and superior tomato varieties. However, until recently, the identification of causal mutations that underlie particular phenotypes has been a very lengthy task that many laboratories could not afford because of spatial and technical limitations. Here, we describe a simple protocol for identifying causal mutations in tomato using a mapping-by-sequencing strategy. Plants displaying phenotypes of interest are first isolated by screening an EMS mutant collection generated in the miniature cultivar Micro-Tom. A recombinant F2 population is then produced by crossing the mutant with a wild-type (WT; non-mutagenized) genotype, and F2 segregants displaying the same phenotype are subsequently pooled. Finally, whole-genome sequencing and analysis of allele distributions in the pools allow for the identification of the causal mutation. The whole process, from the isolation of the tomato mutant to the identification of the causal mutation, takes 6-12 months. This strategy overcomes many previous limitations, is simple to use and can be applied in most laboratories with limited facilities for plant culture and genotyping.

  14. Reads2Type: a web application for rapid microbial taxonomy identification

    DEFF Research Database (Denmark)

    Saputra, Dhany; Rasmussen, Simon; Larsen, Mette Voldby

    2015-01-01

    Identification of bacteria may be based on sequencing and molecular analysis of a specific locus such as 16S rRNA, or a set of loci such as in multilocus sequence typing. In the near future, healthcare institutions and routine diagnostic microbiology laboratories may need to sequence the entire g......, as the entire computational analysis is done on the computer of whom utilizes the web application. This also prevents data privacy issues to arise. The Reads2Type tool is available at http://www.cbs.dtu.dk/~dhany/reads2type.html .......Identification of bacteria may be based on sequencing and molecular analysis of a specific locus such as 16S rRNA, or a set of loci such as in multilocus sequence typing. In the near future, healthcare institutions and routine diagnostic microbiology laboratories may need to sequence the entire...... genome of microbial isolates. Therefore we have developed Reads2Type, a web-based tool for taxonomy identification based on whole bacterial genome sequence data. Raw sequencing data provided by the user are mapped against a set of marker probes that are derived from currently available bacteria complete...

  15. Rapid plant identification using species- and group-specific primers targeting chloroplast DNA.

    Directory of Open Access Journals (Sweden)

    Corinna Wallinger

    Full Text Available Plant identification is challenging when no morphologically assignable parts are available. There is a lack of broadly applicable methods for identifying plants in this situation, for example when roots grow in mixture and for decayed or semi-digested plant material. These difficulties have also impeded the progress made in ecological disciplines such as soil- and trophic ecology. Here, a PCR-based approach is presented which allows identifying a variety of plant taxa commonly occurring in Central European agricultural land. Based on the trnT-F cpDNA region, PCR assays were developed to identify two plant families (Poaceae and Apiaceae, the genera Trifolium and Plantago, and nine plant species: Achillea millefolium, Fagopyrum esculentum, Lolium perenne, Lupinus angustifolius, Phaseolus coccineus, Sinapis alba, Taraxacum officinale, Triticum aestivum, and Zea mays. These assays allowed identification of plants based on size-specific amplicons ranging from 116 bp to 381 bp. Their specificity and sensitivity was consistently high, enabling the detection of small amounts of plant DNA, for example, in decaying plant material and in the intestine or faeces of herbivores. To increase the efficacy of identifying plant species from large number of samples, specific primers were combined in multiplex PCRs, allowing screening for multiple species within a single reaction. The molecular assays outlined here will be applicable manifold, such as for root- and leaf litter identification, botanical trace evidence, and the analysis of herbivory.

  16. Rapid and Accurate Identification by Real-Time PCR of Biotoxin-Producing Dinoflagellates from the Family Gymnodiniaceae

    Directory of Open Access Journals (Sweden)

    Kirsty F. Smith

    2014-03-01

    Full Text Available The identification of toxin-producing dinoflagellates for monitoring programmes and bio-compound discovery requires considerable taxonomic expertise. It can also be difficult to morphologically differentiate toxic and non-toxic species or strains. Various molecular methods have been used for dinoflagellate identification and detection, and this study describes the development of eight real-time polymerase chain reaction (PCR assays targeting the large subunit ribosomal RNA (LSU rRNA gene of species from the genera Gymnodinium, Karenia, Karlodinium, and Takayama. Assays proved to be highly specific and sensitive, and the assay for G. catenatum was further developed for quantification in response to a bloom in Manukau Harbour, New Zealand. The assay estimated cell densities from environmental samples as low as 0.07 cells per PCR reaction, which equated to three cells per litre. This assay not only enabled conclusive species identification but also detected the presence of cells below the limit of detection for light microscopy. This study demonstrates the usefulness of real-time PCR as a sensitive and rapid molecular technique for the detection and quantification of micro-algae from environmental samples.

  17. Vibrational spectroscopy--a powerful tool for the rapid identification of microbial cells at the single-cell level.

    Science.gov (United States)

    Harz, M; Rösch, P; Popp, J

    2009-02-01

    Rapid microbial detection and identification with a high grade of sensitivity and selectivity is a great and challenging issue in many fields, primarily in clinical diagnosis, pharmaceutical, or food processing technology. The tedious and time-consuming processes of current microbiological approaches call for faster ideally on-line identification techniques. The vibrational spectroscopic techniques IR absorption and Raman spectroscopy are noninvasive methods yielding molecular fingerprint information; thus, allowing for a fast and reliable analysis of complex biological systems such as bacterial or yeast cells. In this short review, we discuss recent vibrational spectroscopic advances in microbial identification of yeast and bacterial cells for bulk environment and single-cell analysis. IR absorption spectroscopy enables a bulk analysis whereas micro-Raman-spectroscopy with excitation in the near infrared or visible range has the potential for the analysis of single bacterial and yeast cells. The inherently weak Raman signal can be increased up to several orders of magnitude by applying Raman signal enhancement methods such as UV-resonance Raman spectroscopy with excitation in the deep UV region, surface enhanced Raman scattering, or tip-enhanced Raman scattering. Copyright 2008 International Society for Advancement of Cytometry

  18. Shell thickness-dependent Raman enhancement for rapid identification and detection of pesticide residues at fruit peels.

    Science.gov (United States)

    Liu, Bianhua; Han, Guangmei; Zhang, Zhongping; Liu, Renyong; Jiang, Changlong; Wang, Suhua; Han, Ming-Yong

    2012-01-03

    Here, we report the shell thickness-dependent Raman enhancement of silver-coated gold nanoparticles (Au@Ag NPs) for the identification and detection of pesticide residues at various fruit peels. The Raman enhancement of Au@Ag NPs to a large family of sulfur-containing pesticides is ~2 orders of magnitude stronger than those of bare Au and Ag NPs, and there is a strong dependence of the Raman enhancement on the Ag shell thickness. It has been shown for the first time that the huge Raman enhancement is contributed by individual Au@Ag NPs rather than aggregated Au@Ag NPs with "hot spots" among the neighboring NPs. Therefore, the Au@Ag NPs with excellent individual-particle enhancement can be exploited as stand-alone-particle Raman amplifiers for the surface identification and detection of pesticide residues at various peels of fruits, such as apple, grape, mango, pear, and peach. By casting the particle sensors onto fruit peels, several types of pesticide residues (e.g., thiocarbamate and organophosphorous compounds) have been reliably/rapidly detected, for example, 1.5 nanograms of thiram per square centimeter at apple peel under the current unoptimized condition. The surface-lifting spectroscopic technique offers great practical potentials for the on-site assessment and identification of pesticide residues in agricultural products. © 2011 American Chemical Society

  19. Rapid identification of salmonella serotypes with stereo and hyperspectral microscope imaging Methods

    Science.gov (United States)

    The hyperspectral microscope imaging (HMI) method can reduce detection time within 8 hours including incubation process. The early and rapid detection with this method in conjunction with the high throughput capabilities makes HMI method a prime candidate for implementation for the food industry. Th...

  20. Rapid Detection & Identification of Bacillus Species using MALDI-TOF/TOF and Biomarker Database

    Science.gov (United States)

    2006-06-01

    Dieckmann, R., Graeber, I., Kaesler, I., Szewzyk, U., and von D6hren, H. (2005). Rapid screening and dereplication of bacterial isolates from marine sponges...Bacteriology, vol. 2. Gram- positive bacteria other than Actinomycetes . 1’ ed. Baltimore: Williams & Wilkins, section 13, p 1104-1139. 18 DRDC Suffield

  1. Rapid identification of heterogeneous mixture components with hyperspectral coherent anti-Stokes Raman scattering imaging

    NARCIS (Netherlands)

    Garbacik, E.T.; Herek, Jennifer Lynn; Otto, Cornelis; Offerhaus, Herman L.

    2012-01-01

    For the rapid analysis of complicated heterogeneous mixtures, we have developed a method to acquire and intuitively display hyperspectral coherent anti-Stokes Raman scattering (CARS) images. The imaging is performed with a conventional optical setup based around an optical parametric oscillator.

  2. Rapid catalase supplemental test for identification of members of the family Enterobacteriaceae.

    OpenAIRE

    Chester, B; Moskowitz, L B

    1987-01-01

    A simple, rapid, semiquantitative slide catalase test useful for differentiating members of the family Enterobacteriaceae is described. Judging by the time required for appearance of oxygen bubbles in 3% hydrogen peroxide, the immediate catalase reactors were Yersinia, Serratia, Proteus, Morganella, Providencia, Cedecea, and Hafnia spp. The delayed catalase reactors were Escherichia, Shigella, Klebsiella, Enterobacter, Salmonella, Citrobacter, Edwardsiella, Kluyvera, and Tatumella spp. This i...

  3. Reagent deposition for rapid multiplex pathogen identification in human blood culture samples

    DEFF Research Database (Denmark)

    Mogensen, Klaus Bo; Machado, Ana Manuel; Dufva, Martin

    2014-01-01

    viewed in a dual-color microscope configuration. The test takes 20-30 min to perform. In order to lower the cost of the test, rapid automated reagent deposition is needed. Here, ultrasonic spray coating of polyvinyl alcohol/PNA-probes on microscope glass slides is presented. Different wetting regimes...

  4. Improved identification and quantitation of mature endogenous peptides in the rodent hypothalamus using a rapid conductive sample heating system.

    Science.gov (United States)

    Yang, Ning; Anapindi, Krishna D B; Romanova, Elena V; Rubakhin, Stanislav S; Sweedler, Jonathan V

    2017-11-03

    Measurement, identification, and quantitation of endogenous peptides in tissue samples by mass spectrometry (MS) contribute to our understanding of the complex molecular mechanisms of numerous biological phenomena. For accurate results, it is essential to arrest the postmortem degradation of ubiquitous proteins in samples prior to performing peptidomic measurements. Doing so ensures that the detection of endogenous peptides, typically present at relatively low levels of abundance, is not overwhelmed by protein degradation products. Heat stabilization has been shown to inactivate the enzymes in tissue samples and minimize the presence of protein degradation products in the subsequent peptide extracts. However, the efficacy of different heat treatments to preserve the integrity of full-length endogenous peptides has not been well documented; prior peptidomic studies of heat stabilization methods have not distinguished between the full-length (mature) and numerous truncated (possible artifacts of sampling) forms of endogenous peptides. We show that thermal sample treatment via rapid conductive heat transfer is effective for detection of mature endogenous peptides in fresh and frozen rodent brain tissues. Freshly isolated tissue processing with the commercial Stabilizor T1 heat stabilization system resulted in the confident identification of 65% more full-length mature neuropeptides compared to widely used sample treatment in a hot water bath. This finding was validated by a follow-up quantitative multiple reaction monitoring MS analysis of select neuropeptides. The rapid conductive heating in partial vacuum provided by the Stabilizor T1 effectively reduces protein degradation and decreases the chemical complexity of the sample, as assessed by determining total protein content. This system enabled the detection, identification, and quantitation of neuropeptides related to 22 prohormones expressed in individual rat hypothalami and suprachiasmatic nuclei.

  5. Automatic and rapid identification of glycopeptides by nano-UPLC-LTQ-FT-MS and proteomic search engine.

    Science.gov (United States)

    Giménez, Estela; Gay, Marina; Vilaseca, Marta

    2017-01-30

    Here we demonstrate the potential of nano-UPLC-LTQ-FT-MS and the Byonic™ proteomic search engine for the separation, detection, and identification of N- and O-glycopeptide glycoforms in standard glycoproteins. The use of a BEH C18 nanoACQUITY column allowed the separation of the glycopeptides present in the glycoprotein digest and a baseline-resolution of the glycoforms of the same glycopeptide on the basis of the number of sialic acids. Moreover, we evaluated several acquisition strategies in order to improve the detection and characterization of glycopeptide glycoforms with the maximum number of identification percentages. The proposed strategy is simple to set up with the technology platforms commonly used in proteomic labs. The method allows the straightforward and rapid obtention of a general glycosylated map of a given protein, including glycosites and their corresponding glycosylated structures. The MS strategy selected in this work, based on a gas phase fractionation approach, led to 136 unique peptides from four standard proteins, which represented 78% of the total number of peptides identified. Moreover, the method does not require an extra glycopeptide enrichment step, thus preventing the bias that this step could cause towards certain glycopeptide species. Data are available via ProteomeXchange with identifier PXD003578. We propose a simple and high-throughput glycoproteomics-based methodology that allows the separation of glycopeptide glycoforms on the basis of the number of sialic acids, and their automatic and rapid identification without prior knowledge of protein glycosites or type and structure of the glycans. Copyright © 2016 Elsevier B.V. All rights reserved.

  6. Poly(4-vinylphenol gate insulator with cross-linking using a rapid low-power microwave induction heating scheme for organic thin-film-transistors

    Directory of Open Access Journals (Sweden)

    Ching-Lin Fan

    2016-03-01

    Full Text Available A Microwave-Induction Heating (MIH scheme is proposed for the poly(4-vinylphenol (PVP gate insulator cross-linking process to replace the traditional oven heating cross-linking process. The cross-linking time is significantly decreased from 1 h to 5 min by heating the metal below the PVP layer using microwave irradiation. The necessary microwave power was substantially reduced to about 50 W by decreasing the chamber pressure. The MIH scheme is a good candidate to replace traditional thermal heating for cross-linking of PVP as the gate insulator for organic thin-film-transistors.

  7. Poly(4-vinylphenol) gate insulator with cross-linking using a rapid low-power microwave induction heating scheme for organic thin-film-transistors

    Science.gov (United States)

    Fan, Ching-Lin; Shang, Ming-Chi; Hsia, Mao-Yuan; Wang, Shea-Jue; Huang, Bohr-Ran; Lee, Win-Der

    2016-03-01

    A Microwave-Induction Heating (MIH) scheme is proposed for the poly(4-vinylphenol) (PVP) gate insulator cross-linking process to replace the traditional oven heating cross-linking process. The cross-linking time is significantly decreased from 1 h to 5 min by heating the metal below the PVP layer using microwave irradiation. The necessary microwave power was substantially reduced to about 50 W by decreasing the chamber pressure. The MIH scheme is a good candidate to replace traditional thermal heating for cross-linking of PVP as the gate insulator for organic thin-film-transistors.

  8. Chromosome-Specific DNA Repeats: Rapid Identification in Silico and Validation Using Fluorescence in Situ Hybridization

    Directory of Open Access Journals (Sweden)

    Heinz-Ulrich G. Weier

    2012-12-01

    Full Text Available Chromosome enumeration in interphase and metaphase cells using fluorescence in situ hybridization (FISH is an established procedure for the rapid and accurate cytogenetic analysis of cell nuclei and polar bodies, the unambiguous gender determination, as well as the definition of tumor-specific signatures. Present bottlenecks in the procedure are a limited number of commercial, non-isotopically labeled probes that can be combined in multiplex FISH assays and the relatively high price and effort to develop additional probes. We describe a streamlined approach for rapid probe definition, synthesis and validation, which is based on the analysis of publicly available DNA sequence information, also known as “database mining”. Examples of probe preparation for the human gonosomes and chromosome 16 as a selected autosome outline the probe selection strategy, define a timeline for expedited probe production and compare this novel selection strategy to more conventional probe cloning protocols.

  9. Identification at the crime scene: The sooner, the better? The interpretation of rapid identification information by CSIs at the crime scene.

    Science.gov (United States)

    de Gruijter, Madeleine; Nee, Claire; de Poot, Christianne J

    2017-07-01

    New technologies will allow Crime Scene Investigators (CSIs) in the near future to analyse traces at the crime scene and receive identification information while still conducting the investigation. These developments could have considerable effects on the way an investigation is conducted. CSIs may start reasoning based on possible database-matches which could influence scenario formation (i.e. the construction of narratives that explain the observed traces) during very early phases of the investigation. The goal of this study is to gain more insight into the influence of the rapid identification information on the reconstruction of the crime and the evaluation of traces by addressing two questions, namely 1) is scenario formation influenced from the moment that ID information is provided and 2) do database matches influence the evaluation of traces and the reconstruction of the crime. We asked 48 CSIs from England to investigate a potential murder crime scene on a computer. Our findings show that the interpretation of the crime scene by CSIs is affected by the moment identification information is provided. This information has a higher influence on scenario formation when provided after an initial scenario has been formed. Also, CSIs seem to attach great value to traces that produce matches with databases and hence yield a name of a known person. Similar traces that did not provide matches were considered less important. We question whether this kind of selective attention is desirable as it may cause ignorance of other relevant information at the crime scene. Copyright © 2017 The Chartered Society of Forensic Sciences. Published by Elsevier B.V. All rights reserved.

  10. Rapid identification of Australian bunyavirus isolates belonging to the Simbu serogroup using indirect ELISA formats.

    Science.gov (United States)

    Blacksell, S D; Lunt, R A; White, J R

    1997-06-01

    The Bunyavirus genus, belonging to the Bunyaviridae family, is comprised of a large group of antigenically and geographically disparate arthropod-borne viruses of medical and veterinary significance. In Australia, viruses belonging to the Simbu serogroup of the Bunyavirus genus, Akabane, Tinaroo, Peaton, Aino, Douglas, Thimiri and Facey's Paddock have been isolated. In this communication we describe two indirect ELISAs, referred to as the Simbu serogroup ELISA (SG-ELISA), and the Simbu typing ELISA (ST-ELISA), for the identification of these Simbu serogroup viruses. Infected cell lysate antigens prepared from Simbu serogroup virus isolates were assessed in the SG-ELISA for reactivity with a mouse monoclonal antibody (4H9/B11/F1). The monoclonal antibody reacted strongly with all Australian members of Simbu serogroup reference viruses and is proposed for use as a serogrouping reagent for Simbu viruses. Furthermore, the ST-ELISA enabled specific identification of viruses from within this group by recognition of characteristic reaction patterns between infected cell lysate antigens and a panel of polyclonal antisera raised to Simbu serogroup viruses.

  11. Rapid identification of a narcotic plant Papaver bracteatum using flow cytometry.

    Science.gov (United States)

    Aragane, Masako; Watanabe, Daisuke; Nakajima, Jun'ichi; Yoshida, Masao; Yoshizawa, Masao; Abe, Tomohiro; Nishiyama, Rei; Suzuki, Jin; Moriyasu, Takako; Nakae, Dai; Sudo, Hiroshi; Sato, Hiroyuki; Hishida, Atuyuki; Kawahara, Nobuo; Makabe, So; Nakamura, Ikuo; Mii, Masahiro

    2014-10-01

    In May 2011, numerous poppy plants closely resembling Papaver bracteatum Lindl., a type of narcotic plant that is illegal in Japan, were distributed directly from several large flower shops or through online shopping throughout Japan, including the Tokyo Metropolitan area. In order to better identify the narcotic plants, the relative nuclear DNA content at the vegetative stage was measured by flow cytometric (FCM) analysis in 3 closely-related species of the genus Papaver section Oxytona, namely P. orientale, P. pseudo-orientale, and P. bracteatum, based on the difference between the chromosome numbers of these species. The results showed that the nuclear DNA content differed between these 3 species, and that most of the commercially distributed plants examined in this study could be identified as P. bracteatum. The remaining plants were P. pseudo-orientale, a non-narcotic plant. In addition, the FCM results for the identification of P. bracteatum completely agreed with the results obtained by the morphological analysis, the inter-genic spacer sequence of rpl16-rpl14 (PS-ID sequence) of chloroplast DNA, and the presence of thebaine. These results clearly indicate the usefulness of FCM analysis for the identification of P. bracteatum plants, including when they are in their vegetative stage.

  12. Challenges to the rapid identification of children who have been trafficked for commercial sexual exploitation.

    Science.gov (United States)

    Rafferty, Yvonne

    2016-02-01

    Child trafficking for commercial sexual exploitation (CSE) is a complex phenomenon, requiring multifaceted programs and policies by various stakeholders. A number of publications have focused on preventing this heinous crime. Less attention, however, has been paid to the recovery and rehabilitation of children who have been traumatized as a result of being trafficked for CSE. This article focuses on the first step in the protection and recovery process, which is to ensure that procedures are in place for their identification, so that they might access timely and appropriate assistance. It highlights three situational and two child-related challenges to identification. In addition, it describes the additional victimization experienced by children who are wrongly arrested for crimes associated with prostitution or illegal border crossings, rather than being identified as victims. An extensive literature review was conducted, and included academic publications, as well as governmental and non-governmental reports. In addition, field-based qualitative research was undertaken in South and Southeast Asia, and involved interviews with representatives from United Nations and governmental agencies, non-governmental organizations (NGOs), and aftercare recovery programs. Copyright © 2015 Elsevier Ltd. All rights reserved.

  13. Application of the antibiotic batumin for accurate and rapid identification of staphylococcal small colony variants

    Directory of Open Access Journals (Sweden)

    Churkina Larisa N

    2012-07-01

    Full Text Available Abstract Background Staphylococcus aureus is a major human pathogen causing significant morbidity and mortality. The S. aureus colonies in osteomyelitis, in patients with cystic fibrosis and patients with endoprosthesis rejection frequently have an atypical morphology, i.e. staphylococcal small-colony variants, which form a naturally occurring subpopulation of clinically important staphylococci. Identification of these small colony variants is difficult, because of the loss of typical phenotypic characteristics of these variants. We wanted to improve and simplify the diagnosis of staphylococcal infection using a diagnostic preparation, consisting of 5 μg batumin paper disks. Batumin possesses a unique selective activity against all studied Staphylococcus spp., whereas all other species tested thus far are batumin resistant. We assessed the efficacy of the batumin diagnostic preparation to identify staphylococcal small colony variants, isolated from osteomyelitis patients. Findings With the batumin diagnostic preparation, all 30 tested staphylococcal small-colony variants had a growth inhibition zone around the disk of minimum 25 mm, accordant with the inhibition zones of the parent strains, isolated from the same patients. Conclusions The batumin diagnostic preparation correctly identified the small-colony variants of S. aureus, S. haemolyticus and S. epidermidis as belonging to the genus Staphylococcus, which differ profoundly from parental strains and are difficult to identify with standard methods. Identification of staphylococcal small-colony variants with the batumin diagnostic preparation is technically simple and can facilitate practical laboratory work.

  14. Rapid identification of Mycobacterium avium clinical isolates by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry.

    Science.gov (United States)

    Lin, Chuan-Sheng; Su, Chih-Cheng; Hsieh, Shang-Chen; Lu, Chia-Chen; Wu, Tsu-Lan; Jia, Ju-Hsin; Wu, Ting-Shu; Han, Chau-Chung; Tsai, Wen-Cherng; Lu, Jang-Jih; Lai, Hsin-Chih

    2015-04-01

    Rapid and accurate discrimination of Mycobacterium avium from other mycobacteria is essential for appropriate therapeutic management and timely intervention for infection control. However, routine clinical identification methods for M. avium are both time consuming and labor intensive. In the present study, matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF MS) was used to identify specific cellular protein pattern for rapid identification of M. avium isolates. A total of 40 clinically relevant Mycobacterium strains comprising 13 distinct species were enrolled for the MALDI-TOF MS identification. A 10-minute extraction-free examination procedure was set up to obtain mass spectral fingerprints from whole bacterial cells. The characteristic mass spectral peak patterns in the m/z (mass/charge ratio) range of 5-20 kDa can be obtained within 10 minutes. The species-specific mass spectra for M. avium is identified and can be differentiated from as Mycobacterium strains. This technique shortens and simplifies the identification procedure of MALDI-TOF MS and may further extend the mycobacterial MALDI-TOF MS database. Simplicity and rapidity of identification procedures make MALDI-TOF MS an attractive platform in routine identification of mycobacteria. MALDI-TOF MS is applicable for rapid discrimination of M. avium from other Mycobacterium species, and shows its potential for clinical application. Copyright © 2013. Published by Elsevier B.V.

  15. Combining non-invasive methods for the rapid assessment of mammalian richness in a transect-quadrat survey scheme - Case Study of the Horsh Ehden Nature Reserve, North Lebanon.

    Science.gov (United States)

    Nader, Manal R; El Indary, Shadi; Abi Salloum, Bachir; Abou Dagher, Manale

    2011-01-01

    Lebanon lacks updated information related to the status of mammalian species, their richness and distribution. This study aimed at developing a rapid assessment methodology combining three non-invasive techniques based on the transect-quadrat survey scheme to measure mammalian richness at the Horsh Ehden Nature Reserve. The achieved results showed that the combination of the three techniques, droppings, footprints and photo-trapping for the rapid assessment of mammalian richness supported by Geographical Information System applications is highly effective. Twenty visits covering twenty quadrats out of 49 over a period of nine months allowed the detection of 12 of the 14 targeted species with droppings providing the majority of evidence while footprints and photo-trapping being of equal efficiency. The method can be easily replicated in any region to rapidly assess mammalian richness and the area of activity of the detected species and therefore directing conservation and management activities towards species of interest.

  16. Rapid label-free identification of mixed bacterial infections by surface plasmon resonance

    Directory of Open Access Journals (Sweden)

    Fu Weiling

    2011-06-01

    Full Text Available Abstract Background Early detection of mixed aerobic-anaerobic infection has been a challenge in clinical practice due to the phenotypic changes in complex environments. Surface plasmon resonance (SPR biosensor is widely used to detect DNA-DNA interaction and offers a sensitive and label-free approach in DNA research. Methods In this study, we developed a single-stranded DNA (ssDNA amplification technique and modified the traditional SPR detection system for rapid and simultaneous detection of mixed infections of four pathogenic microorganisms (Pseudomonas aeruginosa, Staphylococcus aureus, Clostridium tetani and Clostridium perfringens. Results We constructed the circulation detection well to increase the sensitivity and the tandem probe arrays to reduce the non-specific hybridization. The use of 16S rDNA universal primers ensured the amplification of four target nucleic acid sequences simultaneously, and further electrophoresis and sequencing confirmed the high efficiency of this amplification method. No significant signals were detected during the single-base mismatch or non-specific probe hybridization (P 2 values of >0.99. The lowest detection limits were 0.03 nM for P. aeruginosa, 0.02 nM for S. aureus, 0.01 nM for C. tetani and 0.02 nM for C. perfringens. The SPR biosensor had the same detection rate as the traditional culture method (P Conclusions Our method can rapidly and accurately identify the mixed aerobic-anaerobic infection, providing a reliable alternative to bacterial culture for rapid bacteria detection.

  17. Identification of pollutant sources in a rapidly developing urban river catchment in China

    Science.gov (United States)

    Huang, Jingshui; Yin, Hailong; Jomma, Seifeddine; Rode, Michael; Zhou, Qi

    2016-04-01

    Rapid economic development and urbanization worldwide cause serious ecological and environmental problems. A typical region that is in transition and requires systemic research for effective intervention is the rapidly developing city of Hefei in central P. R. China. In order to investigate the sources of pollutants over a one-year period in Nanfei River catchment that drains the city of Hefei, discharges were measured and water samples were taken and measured along the 14km river section at 10 sites for 4 times from 2013 to 2014. Overflow concentrations of combined sewer and separate storm drains were also measured by selecting 15 rain events in 4 typical drainage systems. Loads and budgets of water and different pollutant sources i.e., wastewater treatment plant (WWTP) effluent, urban drainage overflow, unknown wastewater were calculated. The water balance demonstrated that >70% of the discharge originated from WWTP effluent. Lack of clean upstream inflow thereby is threatening ecological safety and water quality. Furthermore, mass fluxes calculations revealed that >40% of the COD (Chemical Oxygen Demand) loads were from urban drainage overflow because of a large amount of discharge of untreated wastewater in pumping stations during rain events. WWTP effluent was the predominant source of the total nitrogen loads (>60%) and ammonia loads (>45%). However, the total phosphorous loads from three different sources are similar (˜1/3). Thus, our research provided a basis for appropriate and prior mitigation strategies (state-of-art of WWTP upgrade, sewer systems modification, storm water regulation and storage capacity improvement, etc.) for different precedence-controlled pollutants with the limited infrastructure investments in these rapidly developing urban regions.

  18. Identification and manipulation of the pleuromutilin gene cluster from Clitopilus passeckerianus for increased rapid antibiotic production

    Science.gov (United States)

    Bailey, Andy M.; Alberti, Fabrizio; Kilaru, Sreedhar; Collins, Catherine M.; de Mattos-Shipley, Kate; Hartley, Amanda J.; Hayes, Patrick; Griffin, Alison; Lazarus, Colin M.; Cox, Russell J.; Willis, Christine L.; O'Dwyer, Karen; Spence, David W.; Foster, Gary D.

    2016-05-01

    Semi-synthetic derivatives of the tricyclic diterpene antibiotic pleuromutilin from the basidiomycete Clitopilus passeckerianus are important in combatting bacterial infections in human and veterinary medicine. These compounds belong to the only new class of antibiotics for human applications, with novel mode of action and lack of cross-resistance, representing a class with great potential. Basidiomycete fungi, being dikaryotic, are not generally amenable to strain improvement. We report identification of the seven-gene pleuromutilin gene cluster and verify that using various targeted approaches aimed at increasing antibiotic production in C. passeckerianus, no improvement in yield was achieved. The seven-gene pleuromutilin cluster was reconstructed within Aspergillus oryzae giving production of pleuromutilin in an ascomycete, with a significant increase (2106%) in production. This is the first gene cluster from a basidiomycete to be successfully expressed in an ascomycete, and paves the way for the exploitation of a metabolically rich but traditionally overlooked group of fungi.

  19. Loop-mediated isothermal amplification (LAMP)-based method for rapid mushroom species identification.

    Science.gov (United States)

    Vaagt, Franziska; Haase, Ilka; Fischer, Markus

    2013-02-27

    Toxic mushroom species, such as the death cap ( Amanita phalloides ), are responsible for most mushroom poisonings. In the present work, novel loop-mediated isothermal amplification (LAMP) assays were used for the differentiation of even closely related edible and toxic mushroom species. The applicability of these methods was tested by cross-reaction studies and analysis of spiked mushroom samples (raw and fried material). Contaminations at the level of 2% (w/w) could be detected in different mushroom blends. Three detection methods were used: agarose gel analysis, fluorimetric real-time detection, and visual detection by lateral flow dipsticks (LFD). The LAMP assay combined with LFD detection allows the identification of A. phalloides in about 2 h (including DNA extraction) at a very low level of technical equipment (micropestle, water bath, and mobile centrifuge), which makes this technique perfectly suited for on-site applications.

  20. Sex differential marker FD for rapid sex identification of Litsea cubeba.

    Science.gov (United States)

    Wu, Q; Chen, Y; Wang, Y; Lin, L

    2015-10-21

    Litsea cubeba is an important economic tree in China. Sex identification of the species is required to reduce breeding costs. Molecular biology is an ideal method to achieve this aim because of the lack of morphological differences between male and female plants, especially at the seedling stage. Sequence-related amplified polymorphism was used to amplify sex-related bands. Following sequencing, the amplified fragment Dwas used to create a sequence-characterized amplified region (SCAR) marker, FD. The SCAR marker is approximately 750 bp, is female-specific, and is expected to be useful for L. cubeba breeding programs. Furthermore, the amplified fragment L had homology to sex-determining-related genes of other species. Quantitative real-time polymerase chain reaction analysis of this fragment during flower bud development identified expression differences between male and female plants.

  1. Rapid identification of illegal synthetic adulterants in herbal anti-diabetic medicines using near infrared spectroscopy

    Science.gov (United States)

    Feng, Yanchun; Lei, Deqing; Hu, Changqin

    We created a rapid detection procedure for identifying herbal medicines illegally adulterated with synthetic drugs using near infrared spectroscopy. This procedure includes a reverse correlation coefficient method (RCCM) and comparison of characteristic peaks. Moreover, we made improvements to the RCCM based on new strategies for threshold settings. Any tested herbal medicine must meet two criteria to be identified with our procedure as adulterated. First, the correlation coefficient between the tested sample and the reference must be greater than the RCCM threshold. Next, the NIR spectrum of the tested sample must contain the same characteristic peaks as the reference. In this study, four pure synthetic anti-diabetic drugs (i.e., metformin, gliclazide, glibenclamide and glimepiride), 174 batches of laboratory samples and 127 batches of herbal anti-diabetic medicines were used to construct and validate the procedure. The accuracy of this procedure was greater than 80%. Our data suggest that this protocol is a rapid screening tool to identify synthetic drug adulterants in herbal medicines on the market.

  2. Rapid Identification of Measles Virus Vaccine Genotype by Real-Time PCR.

    Science.gov (United States)

    Roy, Felicia; Mendoza, Lillian; Hiebert, Joanne; McNall, Rebecca J; Bankamp, Bettina; Connolly, Sarah; Lüdde, Amy; Friedrich, Nicole; Mankertz, Annette; Rota, Paul A; Severini, Alberto

    2017-03-01

    During measles outbreaks, it is important to be able to rapidly distinguish between measles cases and vaccine reactions to avoid unnecessary outbreak response measures such as case isolation and contact investigations. We have developed a real-time reverse transcription-PCR (RT-PCR) method specific for genotype A measles virus (MeV) (MeVA RT-quantitative PCR [RT-qPCR]) that can identify measles vaccine strains rapidly, with high throughput, and without the need for sequencing to determine the genotype. We have evaluated the method independently in three measles reference laboratories using two platforms, the Roche LightCycler 480 system and the Applied Biosystems (ABI) 7500 real-time PCR system. In comparison to the standard real-time RT-PCR method, the MeVA RT-qPCR showed 99.5% specificity for genotype A and 94% sensitivity for both platforms. The new assay was able to detect RNA from five currently used vaccine strains, AIK-C, CAM-70, Edmonston-Zagreb, Moraten, and Shanghai-191. The MeVA RT-qPCR assay has been used successfully for measles surveillance in reference laboratories, and it could be readily deployed to national and subnational laboratories on a wide scale. Copyright © 2017 American Society for Microbiology.

  3. Simple and Rapid Molecular Techniques for Identification of Amylose Levels in Rice Varieties

    Science.gov (United States)

    Cheng, Acga; Ismail, Ismanizan; Osman, Mohamad; Hashim, Habibuddin

    2012-01-01

    The polymorphisms of Waxy (Wx) microsatellite and G-T single-nucleotide polymorphism (SNP) in the Wx gene region were analyzed using simplified techniques in fifteen rice varieties. A rapid and reliable electrophoresis method, MetaPhor agarose gel electrophoresis (MAGE), was effectively employed as an alternative to polyacrylamide gel electrophoresis (PAGE) for separating Wx microsatellite alleles. The amplified products containing the Wx microsatellite ranged from 100 to 130 bp in length. Five Wx microsatellite alleles, namely (CT)10, (CT)11, (CT)16, (CT)17, and (CT)18 were identified. Of these, (CT)11 and (CT)17 were the predominant classes among the tested varieties. All varieties with an apparent amylose content higher than 24% were associated with the shorter repeat alleles; (CT)10 and (CT)11, while varieties with 24% or less amylose were associated with the longer repeat alleles. All varieties with intermediate and high amylose content had the sequence AGGTATA at the 5′-leader intron splice site, while varieties with low amylose content had the sequence AGTTATA. The G-T polymorphism was further verified by the PCR-AccI cleaved amplified polymorphic sequence (CAPS) method, in which only genotypes containing the AGGTATA sequence were cleaved by AccI. Hence, varieties with desirable amylose levels can be developed rapidly using the Wx microsatellite and G-T SNP, along with MAGE. PMID:22754356

  4. Seamless bead to microarray screening: rapid identification of the highest affinity protein ligands from large combinatorial libraries.

    Science.gov (United States)

    Astle, John M; Simpson, Levi S; Huang, Yong; Reddy, M Muralidhar; Wilson, Rosemary; Connell, Steven; Wilson, Johnnie; Kodadek, Thomas

    2010-01-29

    Several approaches have been developed for screening combinatorial libraries or collections of synthetic molecules for agonists or antagonists of protein function, each with its own advantages and limitations. In this report, we describe an experimental platform that seamlessly couples massively parallel bead-based screening of one-bead one-compound combinatorial libraries with microarray-based quantitative comparisons of the binding affinities of the many hits isolated from the bead library. Combined with other technical improvements, this technique allows the rapid identification of the best protein ligands in combinatorial libraries containing millions of compounds without the need for labor-intensive resynthesis of the hits. Copyright (c) 2010 Elsevier Ltd. All rights reserved.

  5. Rapid identification of triterpenoid sulfates and hydroxy fatty acids including two new constituents from Tydemania expeditionis by LC-MS

    Science.gov (United States)

    Zhang, Jian-Long; Kubanek, Julia; Hay, Mark E.; Aalbersberg, William; Ye, Wen-Cai; Jiang, Ren-Wang

    2011-01-01

    Tydemania expeditionis Weber-van Bosse (Udoteaceae) is a weakly calcified green alga. In the present paper, liquid chromatography coupled with photodiode array detection and electrospray mass spectrometry was developed to identify the fingerprint components. A total of four triterpenoid sulfates and three hydroxy fatty acids in the ethyl acetate fraction of the crude extract were structurally characterized on the basis of retention time, online UV spectrum and mass fragmentation pattern. Furthermore, detailed LC-MS analysis revealed two new hydroxy fatty acids, which were then prepared and characterized by extensive NMR analyses. The proposed method provides a scientific and technical platform for the rapid identification of triterpenoid sulfates and hydroxy fatty acids in similar marine algae and terrestrial plants. PMID:21915955

  6. [Rapid Identification of Epicarpium Citri Grandis via Infrared Spectroscopy and Fluorescence Spectrum Imaging Technology Combined with Neural Network].

    Science.gov (United States)

    Pan, Sha-sha; Huang, Fu-rong; Xiao, Chi; Xian, Rui-yi; Ma, Zhi-guo

    2015-10-01

    To explore rapid reliable methods for detection of Epicarpium citri grandis (ECG), the experiment using Fourier Transform Attenuated Total Reflection Infrared Spectroscopy (FTIR/ATR) and Fluorescence Spectrum Imaging Technology combined with Multilayer Perceptron (MLP) Neural Network pattern recognition, for the identification of ECG, and the two methods are compared. Infrared spectra and fluorescence spectral images of 118 samples, 81 ECG and 37 other kinds of ECG, are collected. According to the differences in tspectrum, the spectra data in the 550-1 800 cm(-1) wavenumber range and 400-720 nm wavelength are regarded as the study objects of discriminant analysis. Then principal component analysis (PCA) is applied to reduce the dimension of spectroscopic data of ECG and MLP Neural Network is used in combination to classify them. During the experiment were compared the effects of different methods of data preprocessing on the model: multiplicative scatter correction (MSC), standard normal variable correction (SNV), first-order derivative(FD), second-order derivative(SD) and Savitzky-Golay (SG). The results showed that: after the infrared spectra data via the Savitzky-Golay (SG) pretreatment through the MLP Neural Network with the hidden layer function as sigmoid, we can get the best discrimination of ECG, the correct percent of training set and testing set are both 100%. Using fluorescence spectral imaging technology, corrected by the multiple scattering (MSC) results in the pretreatment is the most ideal. After data preprocessing, the three layers of the MLP Neural Network of the hidden layer function as sigmoid function can get 100% correct percent of training set and 96.7% correct percent of testing set. It was shown that the FTIR/ATR and fluorescent spectral imaging technology combined with MLP Neural Network can be used for the identification study of ECG and has the advantages of rapid, reliable effect.

  7. Rapid Identification and Classification of Listeria spp. and Serotype Assignment of Listeria monocytogenes Using Fourier Transform-Infrared Spectroscopy and Artificial Neural Network Analysis.

    Directory of Open Access Journals (Sweden)

    K F Romanolo

    Full Text Available The use of Fourier Transform-Infrared Spectroscopy (FT-IR in conjunction with Artificial Neural Network software NeuroDeveloper™ was examined for the rapid identification and classification of Listeria species and serotyping of Listeria monocytogenes. A spectral library was created for 245 strains of Listeria spp. to give a biochemical fingerprint from which identification of unknown samples were made. This technology was able to accurately distinguish the Listeria species with 99.03% accuracy. Eleven serotypes of Listeria monocytogenes including 1/2a, 1/2b, and 4b were identified with 96.58% accuracy. In addition, motile and non-motile forms of Listeria were used to create a more robust model for identification. FT-IR coupled with NeuroDeveloper™ appear to be a more accurate and economic choice for rapid identification of pathogenic Listeria spp. than current methods.

  8. Rapid and reliable MALDI-TOF mass spectrometry identification of Candida non-albicans isolates from bloodstream infections.

    Science.gov (United States)

    Pulcrano, Giovanna; Iula, Dora Vita; Vollaro, Antonio; Tucci, Alessandra; Cerullo, Monica; Esposito, Matilde; Rossano, Fabio; Catania, Maria Rosaria

    2013-09-01

    Matrix-assisted laser desorption/ionization time-of-flight (MALDI-TOF) mass spectrometry (MS) fingerprinting has recently become an effective instrument for rapid microbiological diagnostics and in particular for identification of micro-organisms directly in a positive blood culture. The aim of the study was to evaluate a collection of 82 stored yeast isolates from bloodstream infection, by MALDI-TOF MS; 21 isolates were identified also directly from positive blood cultures and in the presence of other co-infecting micro-organisms. Of the 82 isolates grown on plates, 64 (76%) were correctly identified by the Vitek II system and 82 (100%) by MALDI-TOF MS; when the two methods gave different results, the isolate was identified by PCR. MALDI-TOF MS was unreliable in identifying two isolates (Candida glabrata and Candida parapsilosis) directly from blood culture; however, direct analysis from positive blood culture samples was fast and effective for the identification of yeast, which is of great importance for early and adequate treatment. © 2013. Published by Elsevier B.V. All rights reserved.

  9. Duplex real-time PCR assay for rapid identification of Staphylococcus aureus isolates from dairy cow milk.

    Science.gov (United States)

    Pilla, Rachel; Snel, Gustavo G M; Malvisi, Michela; Piccinini, Renata

    2013-05-01

    Staphylococcus aureus isolates from dairy cow mastitis are not always consistent with the characteristic morphology described, and molecular investigation is often needed. The aim of the study was to develop a duplex real-time PCR assay for rapid identification of Staph. aureus isolates, targeting both nuc and Sa442. Overall, 140 isolates collected from dairy cow mastitis in 90 different herds, were tested. All strains had been identified using morphological and biochemical characteristics. DNA from each strain was amplified in real-time PCR assay, to detect nuc or Sa442. Thereafter, a duplex real-time PCR assay was performed, and specificity of the amplified products was assessed by high resolution melting curve analysis. Out of 124 Staph. aureus isolates, 33 did not show the typical morphology or enzymic activity; in 118 strains, the two melt-curve peaks consistent with nuc and Sa442 were revealed, while 2 isolates showed only the peak consistent with Sa442. Four isolates bacteriologically identified as Staph. aureus, were PCR-negative and were further identified as Staph. pseudintermedius by sequencing. Staph. pseudintermedius and coagulase-negative staphylococci did not carry nuc or Sa442. The results showed the correct identification of all isolates, comprehending also coagulase-or nuc-negative Staph. aureus, while other coagulase-positive Staphylococci were correctly identified as non-Staph. aureus. Both sensitivity and specificity were 100%. High resolution melting analysis allowed easy detection of unspecific products. Finally, the duplex real-time PCR was applied directly to 40 milk samples, to detect infected mammary quarters. The assay confirmed the results of bacteriological analysis, on Staph. aureus-positive or-negative samples. Therefore, the proposed duplex real-time PCR could be used in laboratory routine as a cost-effective and powerful tool for high-throughput identification of atypical Staph. aureus isolates causing dairy cow mastitis. Also, it

  10. Rapid and high throughput molecular identification of diverse mosquito species by high resolution melting analysis [version 1; referees: 2 approved

    Directory of Open Access Journals (Sweden)

    Yvonne Ukamaka Ajamma

    2016-08-01

    -species. This approach can be employed for rapid identification of mosquitoes.

  11. Rapid identification of Salmonella in dairy products by P-CEIA

    Directory of Open Access Journals (Sweden)

    Rahman Shokri

    2013-12-01

    Materials and Methods: In order to compare Ab-EIA and P-CEIA, different dilutions of S. typhimorium Ra-30 were prepared in peptone water (BPW, pH=7.4. From the 20 samples of milk and cream, 10 suspicious slamonella contaminated samples had been surveyed by the two methods. Results: The sensitivity of P-CEIA method was 106 cfu/ml while the sensitivity of ab-EIA was estimated to be 105 cfu/ml. The sensitivities of the two methods were equal following heat treatment in the presence of sodium deoxy-cholate. Ab-EIA detected 5 and 3 suspicious samples of milk and cream, respectively. PCEIA detected 3 and 6 of the contaminated samples. The first method required 14 hours less time than the other method.Conclusion: Our data shows that P-CEIA is a rapid, economic and sustainable method for Salmonella detection in dairy products.

  12. Rapid, autonomous analysis of HPGe gamma-ray spectra III: Plutonium identification and characterization

    Energy Technology Data Exchange (ETDEWEB)

    Gosnell, Thomas B. [Lawrence Livermore National Lab. (LLNL), Livermore, CA (United States); Wong, James L. [Lawrence Livermore National Lab. (LLNL), Livermore, CA (United States)

    2014-02-11

    RadID is a gamma-ray spectrum analysis program originally written to assist in the detection of the illicit movement of nuclear material. It is specific to the rapid analysis of HPGe gamma-ray data to reveal the radionuclide signatures of interest that may be present in the spectra. It is an autonomous, rule-based heuristic system that can identify well over 200 radioactive sources in about one second. RadID does not require knowledge of the detector efficiency, the source-todetector distance, or the geometry of the inspected radiation source—including any shielding. In this third of a three-document series we discuss how RadID detects the presence of plutonium isotopes and determines a number of its characteristics, most notably: the spectral characteristics of weapons-grade plutonium, reactor-grade plutonium, and heat-source plutonium used in radioisotope thermoelectric generators.

  13. Rapid identification of genes controlling virulence and immunity in malaria parasites

    KAUST Repository

    Abkallo, Hussein M.

    2017-07-13

    Identifying the genetic determinants of phenotypes that impact disease severity is of fundamental importance for the design of new interventions against malaria. Here we present a rapid genome-wide approach capable of identifying multiple genetic drivers of medically relevant phenotypes within malaria parasites via a single experiment at single gene or allele resolution. In a proof of principle study, we found that a previously undescribed single nucleotide polymorphism in the binding domain of the erythrocyte binding like protein (EBL) conferred a dramatic change in red blood cell invasion in mutant rodent malaria parasites Plasmodium yoelii. In the same experiment, we implicated merozoite surface protein 1 (MSP1) and other polymorphic proteins, as the major targets of strain-specific immunity. Using allelic replacement, we provide functional validation of the substitution in the EBL gene controlling the growth rate in the blood stages of the parasites.

  14. Utilizing web-based geodata for rapid disaster identification and assessment

    Science.gov (United States)

    Leith, Kerry; Schmitt, Michael; Sickert, Salomon; Metzger, Alex; Krautblatter, Michael

    2014-05-01

    Developing methods to rapidly locate and quantify the impact of natural disasters can aid both the coordination of emergency response, and the long-term understanding of natural hazards in a range of environmental settings. Gaining such quantitative data in the aftermath of landslide events is particularly challenging, as the localized nature, steep terrain, and frequent damage to infrastructure caused by common triggering events (e.g. earthquakes, storms) complicates traditional methods of survey and data communication. As a result, the first, and often best overview of disastrous events is typically provided by eyewitness or first-responder photographs distributed through official, or social media networks. Although these images allow for an initial qualitative assessment of the event, their ad-hoc nature does not currently allow for either precise location, or quantitative evaluation of key event parameters (e.g. structural setting, geology, geometry, size, transport path, or total fall height). Here we present two tools designed to facilitate initial location and assessment of key event parameters using a combination of freely available geodata and information derived from eyewitness observations. These tools are currently under development, and rely on the adaptation of existing photogrammetric techniques in order to allow users to rapidly map and quantify event parameters from a combination of ad-hoc media photographs, and existing orthophoto and digital terrain model data (e.g. LiDAR, SRTM, ASTER). By incorporating results in freely-available GIS platforms such as Google Earth, local authorities will be able to to better assess and disseminate information regarding the impact of natural disasters in the critical hours following an event. We expect that quantitative data derived from events will provide important information to allow geohazard researchers to better assess landslide generation, and authorities to better plan responses to future triggering

  15. Development of field-based separations for the rapid identification of uranium and plutonium

    Energy Technology Data Exchange (ETDEWEB)

    Mertz, Carol J.; Kaminski, Michael D.; Shkrob, Ilya A.; Kalensky, Michael; Sullivan, Vivian S.; Tsai, Yifen

    2015-05-08

    The development of rapid, radioanalytical techniques to separate uranium and plutonium from complex, field samples are needed for the timely and accurate determination of nuclear material origin, and processing activities. Widespread use of nuclear power and technology in the world has increased demands on analytical laboratories from the monitoring of numerous low-level, environmental samples with variable compositions. Environmental sampling has proven to be one of the strongest technical measures for detecting nuclear material and activities. With the increase in sampling demands, new technologies must offer improvements such as automation, high throughput, reproducible chemical separations, short analysis times, and reduced costs to be effective. We have been developing a portable, separations system for uranium (U) and plutonium (Pu) separations based upon selective extraction of target elements using an extraction chromatographic resin which would allow for simple and fast identifcation when coupled with the appropriate sample digestor and detection systems. The microfluidic design minimizes elution volumes and concentrates the elements of interest in a purified stream. Flowsheet development and testing was demonstrated on a single, micro-column system with an acidified, iron, uranium, and plutonium nitrate stream. The recovery of Pu was optimized by examining various reducing agents at different concentrations for rapid, quantitative recovery from the flow-through design. Quantitative recovery and high selectivity of U and Pu was achieved in the appropriate stripping stages and provided purified and concentrated U and Pu streams. The microfluidic system suggests automation in a small, footprint unit while exploiting the in-line processing of extraction chromatographic resins as the primary means of concentrating the radionuclides from the raw acidic feed and separating the elements into purified streams.

  16. Enumeration and rapid identification of yeasts during extraction processes of extra virgin olive oil in Tuscany.

    Science.gov (United States)

    Mari, Eleonora; Guerrini, Simona; Granchi, Lisa; Vincenzini, Massimo

    2016-06-01

    The aim of this study was to evaluate the occurrence of yeast populations during different olive oil extraction processes, carried out in three consecutive years in Tuscany (Italy), by analysing crushed pastes, kneaded pastes, oil from decanter and pomaces. The results showed yeast concentrations ranging between 10(3) and 10(5) CFU/g or per mL. Seventeen dominant yeast species were identified by random amplified polymorphic DNA with primer M13 and their identification was confirmed by restriction fragments length polymorphism of ribosomal internal transcribed spacer and sequencing rRNA genes. The isolation frequencies of each species in the collected samples pointed out that the occurrence of the various yeast species in olive oil extraction process was dependent not only on the yeasts contaminating the olives but also on the yeasts colonizing the plant for oil extraction. In fact, eleven dominant yeast species were detected from the washed olives, but only three of them were also found in oil samples at significant isolation frequency. On the contrary, the most abundant species in oil samples, Yamadazyma terventina, did not occur in washed olive samples. These findings suggest a phenomenon of contamination of the plant for oil extraction that selects some yeast species that could affect the quality of olive oil.

  17. Rapid urinary tract infection diagnostics by surface-enhanced Raman spectroscopy (SERS): identification and antibiotic susceptibilities.

    Science.gov (United States)

    Premasiri, W R; Chen, Ying; Williamson, P M; Bandarage, D C; Pyles, C; Ziegler, L D

    2017-04-01

    SERS spectra of 12 bacterial strains of urinary tract infection (UTI) clinical isolates grown and enriched from urine are reported. A partial least squares-discriminant analysis (PLS-DA) classification treatment of these SERS spectra results in strain level identification with >95% sensitivity and >99% specificity. The classification model successfully identified the SERS spectra of a urine-cultured strain not used to build this statistical model. Enrichment was accomplished by a filtration and centrifugation protocol. The predetermined drug susceptibility profiles of these clinical isolates thus allowed the SERS methodology to provide appropriate UTI antibiotic information in less than 1 h. Most of this time was used for sample preparation procedures (enrichment and washing) for this proof of principle study. SERS spectra of the enriched bacterial samples are dominated by nucleotide degradation metabolites: adenine, hypoxanthine, xanthine, guanine, uric acid, AMP, and guanosine. Strain-specific specificity is due to the different relative amounts of these purines contributing to the corresponding SERS spectra of these clinical isolates. All measurements were made at the minimal bacterial concentration in urine for UTI diagnosis (105 cfu/mL). Graphical abstract The relative contribution of each of the seven purines found to contribute to the bacterial SERS spectra are summarized in this bar graph. Although strain specific differences are evident, it can be see how the pattern of contributing purines is more different between the four species than between strains of a given species.

  18. Rapid molecular identification of armored scale insects (Hemiptera: Diaspididae) on Mexican 'Hass' avocado.

    Science.gov (United States)

    Rugman-Jones, Paul F; Morse, Joseph G; Stouthamer, Richard

    2009-10-01

    'Hass' avocado, Persea americana Mill., fruit imported into California from Mexico are infested with high levels of armored scale insects (Hemiptera: Diaspididae), constituting several species. The paucity and delicate nature of morphological characters traditionally used to diagnose armored scales often require careful preparation of slide-mounted specimens and expert knowledge of the group, for their accurate identification. Here, we present a simple, quick, and accurate means to identify armored scales on Mexican avocados, based on amplification of the internal transcribed spacer two of ribosomal DNA, by using the polymerase chain reaction (PCR). This region seems to show a high level of intraspecific conformity among scale specimens originating from different localities. A suite of species-specific reverse PCR primers are combined in a single reaction, with a universal forward primer, and produce a PCR product of a unique size, that after standard gel electrophoresis, allows the direct diagnosis of six diaspidid species: Abgrallaspis aguacatae Evans, Watson & Miller; Hemiberlesia lataniae (Signoret); Hemiberlesia sp. near latania; Hemiberlesia rapax (Comstock); Acutaspis albopicta (Cockerell); and Pinnaspis strachani (Cooley). Two additional species, Diaspis miranda (Cockerell) and Diaspis sp. near miranda, also are separated from the others by using this method and are subsequently diagnosed by secondary digestion of the PCR product with the restriction endonuclease smaI.

  19. Rapid and reliable identification of Gram-negative bacteria and Gram-positive cocci by deposition of bacteria harvested from blood cultures onto the MALDI-TOF plate.

    Science.gov (United States)

    Barnini, Simona; Ghelardi, Emilia; Brucculeri, Veronica; Morici, Paola; Lupetti, Antonella

    2015-06-18

    Rapid identification of the causative agent(s) of bloodstream infections using the matrix-assisted laser desorption/ionization time-of-flight (MALDI-TOF) methodology can lead to increased empirical antimicrobial therapy appropriateness. Herein, we aimed at establishing an easier and simpler method, further referred to as the direct method, using bacteria harvested by serum separator tubes from positive blood cultures and placed onto the polished steel target plate for rapid identification by MALDI-TOF. The results by the direct method were compared with those obtained by MALDI-TOF on bacteria isolated on solid media. Identification of Gram-negative bacilli was 100 % concordant using the direct method or MALDI-TOF on isolated bacteria (96 % with score > 2.0). These two methods were 90 % concordant on Gram-positive cocci (32 % with score > 2.0). Identification by the SepsiTyper method of Gram-positive cocci gave concordant results with MALDI-TOF on isolated bacteria in 87 % of cases (37 % with score > 2.0). The direct method herein developed allows rapid identification (within 30 min) of Gram-negative bacteria and Gram-positive cocci from positive blood cultures and can be used to rapidly report reliable and accurate results, without requiring skilled personnel or the use of expensive kits.

  20. Rapid identification of carbapenemase genes in gram-negative bacteria with an oligonucleotide microarray-based assay.

    Directory of Open Access Journals (Sweden)

    Sascha D Braun

    Full Text Available Rapid molecular identification of carbapenemase genes in Gram-negative bacteria is crucial for infection control and prevention, surveillance and for epidemiological purposes. Furthermore, it may have a significant impact upon determining the appropriate initial treatment and greatly benefit for critically ill patients. A novel oligonucleotide microarray-based assay was developed to simultaneously detect genes encoding clinically important carbapenemases as well as selected extended (ESBL and narrow spectrum (NSBL beta-lactamases directly from clonal culture material within few hours. Additionally, a panel of species specific markers was included to identify Escherichia coli, Pseudomonas aeruginosa, Citrobacter freundii/braakii, Klebsiella pneumoniae and Acinetobacter baumannii. The assay was tested using a panel of 117 isolates collected from urinary, blood and stool samples. For these isolates, phenotypic identifications and susceptibility tests were available. An independent detection of carbapenemase, ESBL and NSBL genes was carried out by various external reference laboratories using PCR methods. In direct comparison, the microarray correctly identified 98.2% of the covered carbapenemase genes. This included blaVIM (13 out of 13, blaGIM (2/2, blaKPC (27/27, blaNDM (5/5, blaIMP-2/4/7/8/13/14/15/16/31 (10/10, blaOXA-23 (12/13, blaOXA-40-group (7/7, blaOXA-48-group (32/33, blaOXA-51 (1/1 and blaOXA-58 (1/1. Furthermore, the test correctly identified additional beta-lactamases [blaOXA-1 (16/16, blaOXA-2 (4/4, blaOXA-9 (33/33, OXA-10 (3/3, blaOXA-51 (25/25, blaOXA-58 (2/2, CTX-M1/M15 (17/17 and blaVIM (1/1]. In direct comparison to phenotypical identification obtained by VITEK or MALDI-TOF systems, 114 of 117 (97.4% isolates, including Acinetobacter baumannii (28/28, Enterobacter spec. (5/5, Escherichia coli (4/4, Klebsiella pneumoniae (62/63, Klebsiella oxytoca (0/2, Pseudomonas aeruginosa (12/12, Citrobacter freundii (1/1 and Citrobacter

  1. Rapid identification of Campylobacter jejuni from poultry carcasses and slaughtering environment samples by real-time PCR.

    Science.gov (United States)

    Ivanova, Mirena; Singh, Randhir; Dharmasena, Muthu; Gong, Chao; Krastanov, Albert; Jiang, Xiuping

    2014-06-01

    The objective of this study was to develop a real-time PCR assay for rapid identification of Campylobacter jejuni and to apply the method in analyzing samples from poultry processing. A C. jejuni-specific primer set targeting a portion of the C. jejuni hippuricase gene was developed. The specificity of the newly designed primer pair was verified using 5 C. jejuni strains and 20 other bacterial strains. Sensitivity was determined to be as low as 1 genome copy per reaction. A total of 73 samples were collected at different sites along the processing line during 2 visits to a poultry slaughterhouse and were examined by direct plating onto modified charcoal cefoperazone deoxycholate agar or after enrichment in Bolton broth followed by plating on modified charcoal cefoperazone deoxycholate agar. The newly developed real-time PCR assay was used to identify the presumptive colonies as belonging to C. jejuni. A real-time PCR assay targeting 16S ribosomal RNA was also applied to determine Campylobacter spp. prevalence. Results from the real-time PCR analysis indicated considerable variability in Campylobacter contamination, with incidence rates of 72.7 and 27.6% for sampling days A and B, respectively. Campylobacter was isolated from 100% of prescalded and preeviscerated carcasses on sampling day A. In contrast, on sampling day B, the highest number of Campylobacter-positive carcasses was recovered after evisceration (60%). The chilling process significantly reduced (P Campylobacter population, but the percentage of positive samples on sampling day A increased to 80%. All samples collected from the processing environment, except scalding tank 3 and the prechiller and chiller tanks, were 100% positive on day A, whereas no campylobacters were isolated from machinery on sampling day B. Our results revealed the widespread of C. jejuni in poultry processing and proved that the newly developed real-time PCR assay is a simple, specific, and inexpensive method for rapid C. jejuni

  2. Direct analysis in real time mass spectrometry for the rapid identification of four highly hazardous pesticides in agrochemicals.

    Science.gov (United States)

    Wang, Lei; Zhao, Pengyue; Zhang, Fengzu; Li, Yanjie; Pan, Canping

    2012-08-30

    Direct analysis in real time (DART) is a new ion source technique, which is conducted in the open air under ambient conditions, applied to the rapid and direct analysis of any material (gases, liquids, and solids) with minimal or no sample preparation. In order to take advantage of the capacity of DART mass spectrometry for the real-time analysis of hazardous ingredients in commercial agrochemicals, a pilot study of rapid qualitative determination of hazardous pesticides was performed. Highly hazardous pesticides were identified by DART ionization coupled to a single-quadrupole mass spectrometer (DART-MS). Acetonitrile was chosen for dissolving samples prior to the analysis. Samples were analyzed by this technique in as little as 5 s. Phorate, carbofuran, ethoprophos and fipronil were be detected directly from commercial agrochemicals. The ionization-related parameters (DART temperature, grid voltage and MS fragment) of these compounds were optimized to obtain highly response. Isotope patterns were taken into consideration for qualitative identification. Relative standard deviations (RSDs, n = 5) of 2.3-15.0% were obtained by measuring the relative abundance of selected isotopes. This study showed that DART-MS technology was able to qualitatively determine the existence of highly hazardous pesticides in commercial pesticide formulations. It is suggested that this technology should be applied for routine monitoring in the market. Copyright © 2012 John Wiley & Sons, Ltd.

  3. Rapid sex identification of papaya (Carica papaya) using multiplex loop-mediated isothermal amplification (mLAMP).

    Science.gov (United States)

    Hsu, Te-Hua; Gwo, Jin-Chywan; Lin, Kuan-Hung

    2012-10-01

    Papaya (Carica papaya L.) is established as a cash crop throughout the tropical and subtropical regions due to its easy adaptation to diverse agricultural conditions, high yields, and prompt returns. The sex types of papaya plants are hermaphrodite, male, and female. Among them, hermaphroditic plants are the major type in papaya production, because the fruit has commercial advantages over that of the other sexes. Sex inheritance in papaya is determined by the M and M(h) dominant alleles in males and hermaphrodites, respectively, and a recessive m allele in females. Currently, all hermaphrodite seeds are not available due to the lethality of dominant homozygosity. Therefore, in this study, six male-hermaphrodite-specific markers were developed for a rapid sex identification using multiplex loop-mediated isothermal amplification (mLAMP) to efficiently and precisely select hermaphroditic individuals in the seedling or early growth stage. The LM1-LAMP assay consisted of two sex-LAMP reactions for amplifying two male-specific markers (T12 and Cpsm90) in one reaction, and showed several advantages in terms of a rapid reaction time (papaya production.

  4. Rapid High-throughput Species Identification of Botanical Material Using Direct Analysis in Real Time High Resolution Mass Spectrometry.

    Science.gov (United States)

    Lesiak, Ashton D; Musah, Rabi A

    2016-10-02

    We demonstrate that direct analysis in real time-high resolution mass spectrometry can be used to produce mass spectral profiles of botanical material, and that these chemical fingerprints can be used for plant species identification. The mass spectral data can be acquired rapidly and in a high throughput manner without the need for sample extraction, derivatization or pH adjustment steps. The use of this technique bypasses challenges presented by more conventional techniques including lengthy chromatography analysis times and resource intensive methods. The high throughput capabilities of the direct analysis in real time-high resolution mass spectrometry protocol, coupled with multivariate statistical analysis processing of the data, provide not only class characterization of plants, but also yield species and varietal information. Here, the technique is demonstrated with two psychoactive plant products, Mitragyna speciosa (Kratom) and Datura (Jimsonweed), which were subjected to direct analysis in real time-high resolution mass spectrometry followed by statistical analysis processing of the mass spectral data. The application of these tools in tandem enabled the plant materials to be rapidly identified at the level of variety and species.

  5. Accurate and rapid identification of the Burkholderia pseudomallei near-neighbour, Burkholderia ubonensis, using real-time PCR.

    Science.gov (United States)

    Price, Erin P; Sarovich, Derek S; Webb, Jessica R; Ginther, Jennifer L; Mayo, Mark; Cook, James M; Seymour, Meagan L; Kaestli, Mirjam; Theobald, Vanessa; Hall, Carina M; Busch, Joseph D; Foster, Jeffrey T; Keim, Paul; Wagner, David M; Tuanyok, Apichai; Pearson, Talima; Currie, Bart J

    2013-01-01

    Burkholderia ubonensis is an environmental bacterium belonging to the Burkholderia cepacia complex (Bcc), a group of genetically related organisms that are associated with opportunistic but generally nonfatal infections in healthy individuals. In contrast, the near-neighbour species Burkholderia pseudomallei causes melioidosis, a disease that can be fatal in up to 95% of cases if left untreated. B. ubonensis is frequently misidentified as B. pseudomallei from soil samples using selective culturing on Ashdown's medium, reflecting both the shared environmental niche and morphological similarities of these species. Additionally, B. ubonensis shows potential as an important biocontrol agent in B. pseudomallei-endemic regions as certain strains possess antagonistic properties towards B. pseudomallei. Current methods for characterising B. ubonensis are laborious, time-consuming and costly, and as such this bacterium remains poorly studied. The aim of our study was to develop a rapid and inexpensive real-time PCR-based assay specific for B. ubonensis. We demonstrate that a novel B. ubonensis-specific assay, Bu550, accurately differentiates B. ubonensis from B. pseudomallei and other species that grow on selective Ashdown's agar. We anticipate that Bu550 will catalyse research on B. ubonensis by enabling rapid identification of this organism from Ashdown's-positive colonies that are not B. pseudomallei.

  6. Accurate and rapid identification of the Burkholderia pseudomallei near-neighbour, Burkholderia ubonensis, using real-time PCR.

    Directory of Open Access Journals (Sweden)

    Erin P Price

    Full Text Available Burkholderia ubonensis is an environmental bacterium belonging to the Burkholderia cepacia complex (Bcc, a group of genetically related organisms that are associated with opportunistic but generally nonfatal infections in healthy individuals. In contrast, the near-neighbour species Burkholderia pseudomallei causes melioidosis, a disease that can be fatal in up to 95% of cases if left untreated. B. ubonensis is frequently misidentified as B. pseudomallei from soil samples using selective culturing on Ashdown's medium, reflecting both the shared environmental niche and morphological similarities of these species. Additionally, B. ubonensis shows potential as an important biocontrol agent in B. pseudomallei-endemic regions as certain strains possess antagonistic properties towards B. pseudomallei. Current methods for characterising B. ubonensis are laborious, time-consuming and costly, and as such this bacterium remains poorly studied. The aim of our study was to develop a rapid and inexpensive real-time PCR-based assay specific for B. ubonensis. We demonstrate that a novel B. ubonensis-specific assay, Bu550, accurately differentiates B. ubonensis from B. pseudomallei and other species that grow on selective Ashdown's agar. We anticipate that Bu550 will catalyse research on B. ubonensis by enabling rapid identification of this organism from Ashdown's-positive colonies that are not B. pseudomallei.

  7. Rapid microsatellite identification from Illumina paired-end genomic sequencing in two birds and a snake.

    Directory of Open Access Journals (Sweden)

    Todd A Castoe

    Full Text Available Identification of microsatellites, or simple sequence repeats (SSRs, can be a time-consuming and costly investment requiring enrichment, cloning, and sequencing of candidate loci. Recently, however, high throughput sequencing (with or without prior enrichment for specific SSR loci has been utilized to identify SSR loci. The direct "Seq-to-SSR" approach has an advantage over enrichment-based strategies in that it does not require a priori selection of particular motifs, or prior knowledge of genomic SSR content. It has been more expensive per SSR locus recovered, however, particularly for genomes with few SSR loci, such as bird genomes. The longer but relatively more expensive 454 reads have been preferred over less expensive Illumina reads. Here, we use Illumina paired-end sequence data to identify potentially amplifiable SSR loci (PALs from a snake (the Burmese python, Python molurus bivittatus, and directly compare these results to those from 454 data. We also compare the python results to results from Illumina sequencing of two bird genomes (Gunnison Sage-grouse, Centrocercus minimus, and Clark's Nutcracker, Nucifraga columbiana, which have considerably fewer SSRs than the python. We show that direct Illumina Seq-to-SSR can identify and characterize thousands of potentially amplifiable SSR loci for as little as $10 per sample--a fraction of the cost of 454 sequencing. Given that Illumina Seq-to-SSR is effective, inexpensive, and reliable even for species such as birds that have few SSR loci, it seems that there are now few situations for which prior hybridization is justifiable.

  8. Rapid identification information and its influence on the perceived clues at a crime scene: An experimental study.

    Science.gov (United States)

    de Gruijter, Madeleine; Nee, Claire; de Poot, Christianne J

    2017-11-01

    Crime scenes can always be explained in multiple ways. Traces alone do not provide enough information to infer a whole series of events that has taken place; they only provide clues for these inferences. CSIs need additional information to be able to interpret observed traces. In the near future, a new source of information that could help to interpret a crime scene and testing hypotheses will become available with the advent of rapid identification techniques. A previous study with CSIs demonstrated that this information had an influence on the interpretation of the crime scene, yet it is still unknown what exact information was used for this interpretation and for the construction of their scenario. The present study builds on this study and gains more insight into (1) the exact investigative and forensic information that was used by CSIs to construct their scenario, (2) the inferences drawn from this information, and (3) the kind of evidence that was selected at the crime scene to (dis)prove this scenario. We asked 48 CSIs to investigate a potential murder crime scene on the computer and explicate what information they used to construct a scenario and to select traces for analysis. The results show that the introduction of rapid ID information at the start of an investigation contributes to the recognition of different clues at the crime scene, but also to different interpretations of identical information, depending on the kind of information available and the scenario one has in mind. Furthermore, not all relevant traces were recognized, showing that important information can be missed during the investigation. In this study, accurate crime scenarios where mainly build with forensic information, but we should be aware of the fact that crime scenes are always contaminated with unrelated traces and thus be cautious of the power of rapid ID at the crime scene. Copyright © 2017 The Chartered Society of Forensic Sciences. Published by Elsevier B.V. All rights

  9. Rapid identification of Salmonella serotypes through hyperspectral microscopy with different lighting sources

    Directory of Open Access Journals (Sweden)

    Matthew Eady

    2016-11-01

    Full Text Available The rapid detection of food-borne pathogenic bacteria is critical to the food industry for preventing the introduction of contaminated product into the marketplace and limiting the spread of outbreaks. Hyperspectral microscope images (HMI are a form of optical detection, which classify bacteria by combining microscope images with a spectrophotometer. The objective of this study was to compare the spectra generated from dark-field HMIs of five live Salmonella serotypes from two lighting sources, metal halide (MH and tungsten halogen (TH, assessing classification accuracy and robustness, between 450 nm and 800 nm. It was found that the MH spectra could be reduced to as few as 10 optimal bands between 594 nm and 630 nm, but TH band reduction decreased accuracy, due to the inherent broader peak structure generated by the TH light source. Collection of HMIs from the two light sources comparing the same cells shows slight differences in scatter intensity patterns. Principal component linear discriminate analysis classified serotype subsets (n = 1800, reporting both MH and TH accuracies at 100%, while the reduced key MH bands achieved 99.4–100% accuracy. Principal component regression calculated the root mean squared error of cross-validation 0.948 for both full spectrum lamps. MH or TH lamps can be effectively used for discriminating bacteria HMIs on a cellular level by serotype, but reducing TH bands may lose crucial classification information.

  10. Rapid Identification of Potential Drugs for Diabetic Nephropathy Using Whole-Genome Expression Profiles of Glomeruli

    Directory of Open Access Journals (Sweden)

    Jingsong Shi

    2016-01-01

    Full Text Available Objective. To investigate potential drugs for diabetic nephropathy (DN using whole-genome expression profiles and the Connectivity Map (CMAP. Methodology. Eighteen Chinese Han DN patients and six normal controls were included in this study. Whole-genome expression profiles of microdissected glomeruli were measured using the Affymetrix human U133 plus 2.0 chip. Differentially expressed genes (DEGs between late stage and early stage DN samples and the CMAP database were used to identify potential drugs for DN using bioinformatics methods. Results. (1 A total of 1065 DEGs (FDR 1.5 were found in late stage DN patients compared with early stage DN patients. (2 Piperlongumine, 15d-PGJ2 (15-delta prostaglandin J2, vorinostat, and trichostatin A were predicted to be the most promising potential drugs for DN, acting as NF-κB inhibitors, histone deacetylase inhibitors (HDACIs, PI3K pathway inhibitors, or PPARγ agonists, respectively. Conclusion. Using whole-genome expression profiles and the CMAP database, we rapidly predicted potential DN drugs, and therapeutic potential was confirmed by previously published studies. Animal experiments and clinical trials are needed to confirm both the safety and efficacy of these drugs in the treatment of DN.

  11. Low-Density Macroarray for Rapid Detection and Identification of Crimean-Congo Hemorrhagic Fever Virus▿

    Science.gov (United States)

    Wölfel, Roman; Paweska, Janusz T.; Petersen, Nadine; Grobbelaar, Antoinette A.; Leman, Patricia A.; Hewson, Roger; Georges-Courbot, Marie-Claude; Papa, Anna; Heiser, Volker; Panning, Marcus; Günther, Stephan; Drosten, Christian

    2009-01-01

    Crimean-Congo hemorrhagic fever (CCHF) is a tick-borne viral zoonosis which occurs throughout Africa, Eastern Europe, and Asia and results in an approximately 30% fatality rate. A reverse transcription-PCR assay including a competitive internal control was developed on the basis of the most up-to-date genome information. Biotinylated amplification products were hybridized to DNA macroarrays on the surfaces of polymer supports, and hybridization events were visualized by incubation with a streptavidin-horseradish peroxidase conjugate and the formation of a visible substrate precipitate. Optimal assay conditions for the detection of as few as 6.3 genome copies per reaction were established. Eighteen geographically and historically diverse CCHF virus strains representing all clinically relevant isolates were detected. The feasibility of the assay for clinical diagnosis was validated with acute-phase patient samples from South Africa, Iran, and Pakistan. The assay provides a specific, sensitive, and rapid method for CCHF virus detection without requiring sophisticated equipment. It has usefulness for the clinical diagnosis and surveillance of CCHF infections under limited laboratory conditions in developing countries or in field situations. PMID:19225100

  12. Low-density macroarray for rapid detection and identification of Crimean-Congo hemorrhagic fever virus.

    Science.gov (United States)

    Wölfel, Roman; Paweska, Janusz T; Petersen, Nadine; Grobbelaar, Antoinette A; Leman, Patricia A; Hewson, Roger; Georges-Courbot, Marie-Claude; Papa, Anna; Heiser, Volker; Panning, Marcus; Günther, Stephan; Drosten, Christian

    2009-04-01

    Crimean-Congo hemorrhagic fever (CCHF) is a tick-borne viral zoonosis which occurs throughout Africa, Eastern Europe, and Asia and results in an approximately 30% fatality rate. A reverse transcription-PCR assay including a competitive internal control was developed on the basis of the most up-to-date genome information. Biotinylated amplification products were hybridized to DNA macroarrays on the surfaces of polymer supports, and hybridization events were visualized by incubation with a streptavidin-horseradish peroxidase conjugate and the formation of a visible substrate precipitate. Optimal assay conditions for the detection of as few as 6.3 genome copies per reaction were established. Eighteen geographically and historically diverse CCHF virus strains representing all clinically relevant isolates were detected. The feasibility of the assay for clinical diagnosis was validated with acute-phase patient samples from South Africa, Iran, and Pakistan. The assay provides a specific, sensitive, and rapid method for CCHF virus detection without requiring sophisticated equipment. It has usefulness for the clinical diagnosis and surveillance of CCHF infections under limited laboratory conditions in developing countries or in field situations.

  13. Rapid identification of mRNA processing defects with a novel single-cell yeast reporter.

    Science.gov (United States)

    Sorenson, Matthew R; Stevens, Scott W

    2014-05-01

    It has become increasingly evident that gene expression processes in eukaryotes involve communication and coordination between many complex, independent macromolecular machines. To query these processes and to explore the potential relationships between them in the budding yeast Saccharomyces cerevisiae, we designed a versatile reporter using multicolor high-throughput flow cytometry. Due to its design, this single reporter exhibits a distinctive signature for many defects in gene expression including transcription, histone modification, pre-mRNA splicing, mRNA export, nonsense-mediated decay, and mRNA degradation. Analysis of the reporter in 4967 nonessential yeast genes revealed striking phenotypic overlaps between chromatin remodeling, histone modification, and pre-mRNA splicing. Additionally, we developed a copper-inducible reporter, with which we demonstrate that 5-fluorouracil mimics the mRNA decay phenotype of cells lacking the 3'-5' exonuclease Rrp6p. Our reporter is capable of performing high-throughput, rapid, and large-scale screens to identify and characterize genetic and chemical perturbations of the major eukaryotic gene expression processes.

  14. CS-SCORE: Rapid identification and removal of human genome contaminants from metagenomic datasets.

    Science.gov (United States)

    Haque, Mohammed Monzoorul; Bose, Tungadri; Dutta, Anirban; Reddy, Chennareddy Venkata Siva Kumar; Mande, Sharmila S

    2015-08-01

    Metagenomic sequencing data, obtained from host-associated microbial communities, are usually contaminated with host genome sequence fragments. Prior to performing any downstream analyses, it is necessary to identify and remove such contaminating sequence fragments. The time and memory requirements of available host-contamination detection techniques are enormous. Thus, processing of large metagenomic datasets is a challenging task. This study presents CS-SCORE--a novel algorithm that can rapidly identify host sequences contaminating metagenomic datasets. Validation results indicate that CS-SCORE is 2-6 times faster than the current state-of-the-art methods. Furthermore, the memory footprint of CS-SCORE is in the range of 2-2.5GB, which is significantly lower than other available tools. CS-SCORE achieves this efficiency by incorporating (1) a heuristic pre-filtering mechanism and (2) a directed-mapping approach that utilizes a novel sequence composition metric (cs-score). CS-SCORE is expected to be a handy 'pre-processing' utility for researchers analyzing metagenomic datasets. For academic users, an implementation of CS-SCORE is freely available at: http://metagenomics.atc.tcs.com/cs-score (or) https://metagenomics.atc.tcs.com/preprocessing/cs-score. Copyright © 2015 Elsevier Inc. All rights reserved.

  15. Rapid identification of vibrio-cholerae O1 by coaglutination test using mono-specifis antibody

    Directory of Open Access Journals (Sweden)

    Bazargan SA

    1996-07-01

    Full Text Available In our investigation, rabbit hyper-immune serum to V.cholerae ogawa was absorbed with V.cholerae inaba whole-cells and vice versa. Applying ammonium sulphate precipitation method, mono-specific g globulins were purified and concentrated from the absorbed whole serum. These antibodies were fixed on staphylococcus cowan 1 NCTC-8325 whole-cells, using different chemical fixatives. It was observed that maximum fixation of g globulin to protein-A was achieved by 1-propanol 50% at 3 hours, which revealed through single radial immuno-diffusion techniqe. The rectal swab samples were cultured in an enrichment bile-peptons broth. After 5 hours 37°C while agitations, one drop of each sample was mixed with one drop of vibrio-cholerae bivalent mono-specific coagglutination reagent (VBCR. The results were read after 2 to 3 minutes. Finally though statistical analysis sensitivity and specificity of coagglutination test were calculated to be 95.1% and 99.2% respectively, when compared to positive & negative controls and conventional culture methods. Using VBCR, coagglutination test can be therefore considered as a simple, reliable and rapid method to detect V.cholerae O1 in the stool of patients in endemic area and less equipped laboratories

  16. Rapid identification and classification of Campylobacter spp. using laser optical scattering technology.

    Science.gov (United States)

    He, Yiping; Reed, Sue; Bhunia, Arun K; Gehring, Andrew; Nguyen, Ly-Huong; Irwin, Peter L

    2015-05-01

    Campylobacter jejuni and Campylobacter coli are the two important species responsible for most of the Campylobacter infections in humans. Reliable isolation and detection of Campylobacter spp. from food samples are challenging due to the interferences from complex food substances and the fastidious growth requirements of this organism. In this study, a novel biosensor-based detection called BARDOT (BActerial Rapid Detection using Optical scattering Technology) was developed for high-throughput screening of Campylobacter colonies grown on an agar plate without disrupting the intact colonies. Image pattern characterization and principal component analysis (PCA) of 6909 bacterial colonies showed that the light scatter patterns of C. jejuni and C. coli were strikingly different from those of Salmonella spp., Escherichia coli O157:H7, and Listeria monocytogenes. Examination of a mixed culture of these microorganisms revealed 85% (34/40) accuracy in differentiating Campylobacter from the other three major foodborne pathogens based on the similarity to the scatter patterns in an established library. The application of BARDOT in real food has been addressed through the analysis of Campylobacter spiked ground chicken and naturally contaminated fresh chicken pieces. Combined with real-time PCR verification, BARDOT was able to identify Campylobacter isolates from retail chicken. Moreover, applying passive filtration to food samples facilitated the isolation of pure Campylobacter colonies and therefore overcame the interference of the food matrix on BARDOT analysis. Published by Elsevier Ltd.

  17. Rapid identification of Chinese Sauce liquor from different fermentation positions with FT-IR spectroscopy

    Science.gov (United States)

    Li, Changwen; Wei, Jiping; Zhou, Qun; Sun, Suqin

    2008-07-01

    FT-IR and two-dimensional correlation spectroscopy (2D-IR) technology were applied to discriminate Chinese Sauce liquor from different fermentation positions (top, middle and bottom of fermentation cellar) for the first time. The liquors at top, middle and bottom of fermentation cellar, possessed the characteristic peaks at 1731 cm -1, 1733 cm -1 and 1602 cm -1, respectively. In the 2D correlation infrared spectra, the differences were amplified. A strong auto-peak at 1725 cm -1 showed in the 2D spectra of the Top Liquor, which indicated that the liquor might contain some ester compounds. Different from Top Liquor, three auto-peaks at 1695, 1590 and 1480 cm -1 were identified in 2D spectra of Middle Liquor, which were the characteristic absorption of acid, lactate. In 2D spectra of Bottom Liquor, two auto-peaks at 1570 and 1485 cm -1 indicated that lactate was the major component. As a result, FT-IR and 2D-IR correlation spectra technology provided a rapid and effective method for the quality analysis of the Sauce liquor.

  18. Rapid identification of the genus Dekkera/Brettanomyces, the Dekkera subgroup and all individual species.

    Science.gov (United States)

    Hulin, M; Harrison, E; Stratford, M; Wheals, A E

    2014-09-18

    The genus Dekkera/Brettanomyces comprises five described species: Dekkera bruxellensis, D. anomala, Brettanomyces custersianus, B. naardenensis and B. nanus. Some of them, especially D. bruxellensis, are important spoilage organisms, particularly in the wine and beverage industries. Because of their economic importance many different methods have been developed to identify members of the genus in general and D. bruxellensis in particular. These methods vary in their rapidity, complexity and cost but, partly because of confidentiality issues, it is unclear which methods are used, or how widely, in the relevant industries. Building on previous work with the genera Saccharomyces and Zygosaccharomyces, a suite of eight PCR primer pairs has been designed either on the D1-D2 region of the 26S rRNA gene or translation elongation factor TEF1-α. These primers can specifically identify the genus as a whole, only Dekkera species, each one of the five recognised species as well as a significant subgroup of D. bruxellensis represented by NCYC 3426. Multiplexing has also been tried and it has been shown to be possible with some combinations of genus or Dekkera-level and species-specific primers. Using direct colony PCR amplification followed by gel electrophoresis, a clear positive result can be obtained in less than 3h, thus providing a quick, reliable and inexpensive way to identify target species. Copyright © 2014 Elsevier B.V. All rights reserved.

  19. A proteomic approach for the rapid, multi-informative and reliable identification of blood.

    Science.gov (United States)

    Patel, E; Cicatiello, P; Deininger, L; Clench, M R; Marino, G; Giardina, P; Langenburg, G; West, A; Marshall, P; Sears, V; Francese, S

    2016-01-07

    Blood evidence is frequently encountered at the scene of violent crimes and can provide valuable intelligence in the forensic investigation of serious offences. Because many of the current enhancement methods used by crime scene investigators are presumptive, the visualisation of blood is not always reliable nor does it bear additional information. In the work presented here, two methods employing a shotgun bottom up proteomic approach for the detection of blood are reported; the developed protocols employ both an in solution digestion method and a recently proposed procedure involving immobilization of trypsin on hydrophobin Vmh2 coated MALDI sample plate. The methods are complementary as whilst one yields more identifiable proteins (as biomolecular signatures), the other is extremely rapid (5 minutes). Additionally, data demonstrate the opportunity to discriminate blood provenance even when two different blood sources are present in a mixture. This approach is also suitable for old bloodstains which had been previously chemically enhanced, as experiments conducted on a 9-year-old bloodstain deposited on a ceramic tile demonstrate.

  20. Rapid identification and enumeration of Saccharomyces cerevisiae cells in wine by real-time PCR.

    Science.gov (United States)

    Martorell, P; Querol, A; Fernández-Espinar, M T

    2005-11-01

    Despite the beneficial role of Saccharomyces cerevisiae in the food industry for food and beverage production, it is able to cause spoilage in wines. We have developed a real-time PCR method to directly detect and quantify this yeast species in wine samples to provide winemakers with a rapid and sensitive method to detect and prevent wine spoilage. Specific primers were designed for S. cerevisiae using the sequence information obtained from a cloned random amplified polymorphic DNA band that differentiated S. cerevisiae from its sibling species Saccharomyces bayanus, Saccharomyces pastorianus, and Saccharomyces paradoxus. The specificity of the primers was demonstrated for typical wine spoilage yeast species. The method was useful for estimating the level of S. cerevisiae directly in sweet wines and red wines without preenrichment when yeast is present in concentrations as low as 3.8 and 5 CFU per ml. This detection limit is in the same order as that obtained from glucose-peptone-yeast growth medium (GPY). Moreover, it was possible to quantify S. cerevisiae in artificially contaminated samples accurately. Limits for accurate quantification in wine were established, from 3.8 x 10(5) to 3.8 CFU/ml in sweet wine and from 5 x 10(6) to 50 CFU/ml in red wine.

  1. Rapid identification of novel antigens of Salmonella Enteritidis by microarray-based immunoscreening.

    Science.gov (United States)

    Danckert, Lena; Hoppe, Sebastian; Bier, Frank F; von Nickisch-Rosenegk, Markus

    2014-01-01

    We report on an approach to rapidly screen thousands of Salmonella Enteritidis proteins with the goal of identifying novel immunodominant proteins. We used a microarray-based system that warrants high throughput and easy handling. Seven immunogenic candidates were selected after screening. Comparative analyses by ELISA and microarrays manifested their immunodominant character. The large repetitive protein (SEN4030) that plays a role as a putative adhesin in initial cell surface interaction and is highly specific to Salmonella is considered to be the most suitable protein for a diagnostic approach. The results further demonstrate that the strategy applied herein is convenient for specifically identifying immunogenic proteins of pathogenic microorganisms. Consequently, it enables a sound assessment of promising candidates for diagnostic applications and vaccine development. Moreover, the elucidation of immunogenic proteins may assist in unveiling unknown virulence-associated factors, thus furthering the understanding of the underlying pathogenicity of Salmonella in general, and of S. Enteritidis, one of the most frequently detected serovars of this pathogen, in particular. FigureThe microarray-based approach was aimed at identifying novel immunodominant proteins of S. Enteritidis. Seven antigens were revealed by screening a cDNA expression library. SEN4030, a large repetitive protein specific for salmonella, is considered an optimal candidate for future applications.

  2. A method for the rapid detection and identification of halo blight pathogen on common bean

    Directory of Open Access Journals (Sweden)

    Popović Tatjana

    2014-01-01

    Full Text Available A diagnostic method based on nested-PCR, followed by ELISA and conventional bacteriology tests, for the rapid and reliable detection of halo blight pathogen Pseudomonas savastanoi pv. phaseolicola (Psp collected from infected bean leaves and seeds is described. Psp formed white, small and flat colonies on nutrient agar medium, creamy white, flat and circular on Milk-Tween agar medium and light yellow, convex and shiny on modified sucrose peptone agar medium. Eighteen Gram-negative, catalase-positive and oxidase-negative strains were subjected to nested PCR with primers P 5.1/P 3.1 and P 5.2/P 3.2, which directed the amplification of the 450 bp target DNA fragment in all tested strains. According to the results of DAS- and PTA-ELISA with respect to reactivity to specific antibodies, all analyzed strains belonged to Psp bacterium. Pathogenicity was tested on bean pods and cotyledon leaves, on which greasy spots were formed. Psp did not cause hypersensitive reaction on the leaves of tobacco and geranium. Strains produced levan, fluorescent pigment, oxidative metabolism of glucose, did not reduce nitrate, did not produce indole and H2S, did not hydrolyze starch, gelatin and esculin; they produced acid from glucose, mannose, sucrose and glycerol, and did not produce acid from maltose, starch, esculin, dulcite, sorbitol, inositol and erythritol.

  3. AOTF-based near-infrared imaging spectrometer for rapid identification of camouflaged target

    Science.gov (United States)

    Gao, Zhifan; Zeng, Libo; Wu, Qiongshui

    2014-11-01

    Acousto-optic tunable filter (AOTF) is a novel device for spectrometer. The electronic tunability qualifies it with the most compelling advantages of higher wavelength scan rate over the conventional spectrometers that are mechanically tuned, and the feature of large angular aperture makes the AOTF particularly suitable in imaging applications. In this research, an AOTF-based near-infrared imaging spectrometer was developed. The spectrometer consists of a TeO2 AOTF module, a near-infrared imaging lens assembly, an AOTF controller, an InGaAs array detector, an image acquisition card, and a PC. A precisely designed optical wedge is placed at the emergent surface of the AOTF to deal with the inherent dispersion of the TeO2 that may degrade the spatial resolution. The direct digital synthesizer (DDS) techniques and the phase locked loop (PLL) techniques are combined for radio frequency (RF) signal synthesis. The PLL is driven by the DDS to take advantage of both their merits of high frequency resolution, high frequency scan rate and strong spurious signals resistance capability. All the functions relating to wavelength scan, image acquisition, processing, storge and display are controlled by the PC. Calibration results indicate that the spectral range is 898~1670 nm, the spectral resolution is 6.8 nm(@1064 nm), the wavelength separation between frames in the spectral image assembly is 1.0 nm, and the processing time of a single image is less than 1 ms if a TV camera with 640×512 detector is incorporated. A prototype device was assembled to test the capability of differentiating samples with similar appearances, and satisfactory results were achieved. By this device, the chemical compositions and the distribution information can be obtained simultaneously. This system has the most advantages of no moving parts, fast wavelength scan and strong vibration resistance. The proposed imaging spectrometer has a significant application prospect in the area of identification of

  4. Rapid identification of human SNAP-25 transcript variants by a miniaturized capillary electrophoresis system.

    Science.gov (United States)

    Németh, Nóra; Kerékgyártó, Márta; Sasvári-Székely, Mária; Rónai, Zsolt; Guttman, András

    2014-02-01

    The 25 kDa synaptosomal-associated protein (SNAP-25) is a crucial component of the soluble N-ethylmaleimide-sensitive factor attachment protein receptor complex and plays an important role in neurotransmission in the central nervous system. SNAP-25 has two different splice variants, SNAP-25a and SNAP-25b, differing in nine amino acids that results in a slight functional alteration of the generated soluble N-ethylmaleimide-sensitive factor attachment protein receptor complex. Two independent techniques, a PCR-miniaturized CE method and a real-time PCR based approach were elaborated for the specific and quantitative detection of the two SNAP-25 transcription variants. DNA-constructs coding for the two isoforms were used for optimization. Excellent specificity was observed with the use of our previously described highly sensitive miniaturized CE system in combination with quantitative PCR. The ratio of the two isoforms were reliably detected in a range of at least four orders of magnitude with a linear regression of R(2) = 0.987. Expression of the two isoforms was determined in human samples, where SNAP-25 was detected even in non-neural tissues, although at approximately a 100-fold lower level compared to the central nervous system. The relative amount of the SNAP-25b isoform was higher in the brain, whereas expression of SNAP-25a variant proved to be slightly higher in extra-neural cell types. The genomics approach in conjunction with the miniaturized CE system introduced in this paper is readily applicable for rapid alternative splice variant analysis. © 2013 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  5. Colour schemes

    DEFF Research Database (Denmark)

    van Leeuwen, Theo

    2013-01-01

    This chapter presents a framework for analysing colour schemes based on a parametric approach that includes not only hue, value and saturation, but also purity, transparency, luminosity, luminescence, lustre, modulation and differentiation.......This chapter presents a framework for analysing colour schemes based on a parametric approach that includes not only hue, value and saturation, but also purity, transparency, luminosity, luminescence, lustre, modulation and differentiation....

  6. Rapid screening and identification of ACE inhibitors in snake venoms using at-line nanofractionation LC-MS.

    Science.gov (United States)

    Mladic, Marija; de Waal, Tessa; Burggraaff, Lindsey; Slagboom, Julien; Somsen, Govert W; Niessen, Wilfried M A; Manjunatha Kini, R; Kool, Jeroen

    2017-10-01

    This study presents an analytical method for the screening of snake venoms for inhibitors of the angiotensin-converting enzyme (ACE) and a strategy for their rapid identification. The method is based on an at-line nanofractionation approach, which combines liquid chromatography (LC), mass spectrometry (MS), and pharmacology in one platform. After initial LC separation of a crude venom, a post-column flow split is introduced enabling parallel MS identification and high-resolution fractionation onto 384-well plates. The plates are subsequently freeze-dried and used in a fluorescence-based ACE activity assay to determine the ability of the nanofractions to inhibit ACE activity. Once the bioactive wells are identified, the parallel MS data reveals the masses corresponding to the activities found. Narrowing down of possible bioactive candidates is provided by comparison of bioactivity profiles after reversed-phase liquid chromatography (RPLC) and after hydrophilic interaction chromatography (HILIC) of a crude venom. Additional nanoLC-MS/MS analysis is performed on the content of the bioactive nanofractions to determine peptide sequences. The method described was optimized, evaluated, and successfully applied for screening of 30 snake venoms for the presence of ACE inhibitors. As a result, two new bioactive peptides were identified: pELWPRPHVPP in Crotalus viridis viridis venom with IC 50  = 1.1 μM and pEWPPWPPRPPIPP in Cerastes cerastes cerastes venom with IC 50  = 3.5 μM. The identified peptides possess a high sequence similarity to other bradykinin-potentiating peptides (BPPs), which are known ACE inhibitors found in snake venoms.

  7. Random amplified ribosomal DNA restriction analysis for rapid identification of thermophilic Actinomycete-like bacteria involved in hypersensitivity pneumonitis.

    Science.gov (United States)

    Harvey, I; Cormier, Y; Beaulieu, C; Akimov, V N; Mériaux, A; Duchaine, C

    2001-07-01

    Hypersensitivity pneumonitis (HP) is a pulmonary disease characterised by inflammation that can be caused by, amongst other substances, a subset of 4 thermophilic mycelial bacteria: Saccharopolyspora rectivirgula, Saccharomonospora viridis, Thermoactinomyces sacchari, and Thermoactinomyces vulgaris. Air sampling analyses in highly contaminated environments are often performed to evaluate exposure to these species which are difficult and fastidious to identify by conventional techniques. The aim of this study was to use amplified ribosomal DNA restriction analysis (ARDRA) to develop a method of identification for those thermophilic organisms that would be more rapid and simple. Strains of these 4 species were obtained from the American type culture collection (ATCC) and were characterized using biochemical tests and ARDRA patterns obtained on their partial-lenght amplified 16S rDNAs. To validate this approach, ARDRA with two restriction enzymes, TaqI and HhaI, was applied to 49 thermophilic actinomycete-like strains from environmental samples (sawmills). The results obtained show that combining some cultural characteristics and biochemical tests, such as xanthine or hypoxanthine decomposition, growth in the presence of NaCl, lysozyme or novobiocin, and spore resistance over 100 degrees C provide a rough identification and selection of the genera of interest. Consequently, target species could be confirmed by digestion of partial-lenght 16S rDNA with the use of Taql and HhaI restriction enzymes that gave specific restriction patterns. ARDRA analyses on the 49 environmental actinomycete-like organisms revealed the presence of 8 Saccharopolyspora rectivirgula, 2 Saccharomonospora viridis, and 15 Thermoactinomyces vulgaris strains, the other strains had restriction patterns different than those of the species of interest. Results of the present study will be applicable to other potential HP environments such as dairy barns, peat bogs and compost plants.

  8. Assessment of impact of peptide nucleic acid fluorescence in situ hybridization for rapid identification of coagulase-negative staphylococci in the absence of antimicrobial stewardship intervention.

    Science.gov (United States)

    Holtzman, Carol; Whitney, Dana; Barlam, Tamar; Miller, Nancy S

    2011-04-01

    Peptide nucleic acid fluorescence in situ hybridization (PNA FISH) was instituted at Boston Medical Center for the rapid identification of coagulase-negative staphylococci (CoNS). Without active notification or antimicrobial stewardship intervention, a pre- and postimpact analysis showed no benefit of this assay with respect to the length of hospital stay or vancomycin use.

  9. Rapid identification and antimicrobial susceptibility testing of positive blood cultures using MALDI-TOF MS and a modification of the standardised disc diffusion test: a pilot study.

    LENUS (Irish Health Repository)

    Fitzgerald, C

    2016-04-27

    In an era when clinical microbiology laboratories are under increasing financial pressure, there is a need for inexpensive, yet effective, rapid microbiology tests. The aim of this study was to evaluate a novel modification of standard methodology for the identification and antimicrobial susceptibility testing (AST) of pathogens in positive blood cultures, reducing the turnaround time of laboratory results by 24 h.

  10. Rapid identification and classification of Listeria spp. and serotype assignment of Listeria monocytogenes using fourier transform-infrared spectroscopy and artificial neural network analysis

    Science.gov (United States)

    The use of Fourier Transform-Infrared Spectroscopy (FT-IR) in conjunction with Artificial Neural Network software, NeuroDeveloper™ was examined for the rapid identification and classification of Listeria species and serotyping of Listeria monocytogenes. A spectral library was created for 245 strains...

  11. Performance of two tube coagulase methods for rapid identification of Staphylococcus aureus from blood cultures and their impact on antimicrobial management.

    NARCIS (Netherlands)

    Sturm, P.D.J.; Kwa, D.; Vos, F.J.; Bartels, C.J.; Schulin, T.

    2008-01-01

    Test parameters and clinical impact of the direct tube coagulase test (DTCT) for rapid identification of Staphylococcus aureus from blood culture were investigated. The sensitivity of the DTCT at 4 h using saline dilution was 96%, compared with 93% using serum separator tubes; specificity was 100%

  12. Development of a triple hyphenated HPLC-radical scavenging detection-DAD-SPE-NMR system for the rapid identification of antioxidants in complex plant extracts

    NARCIS (Netherlands)

    Pukalskas, A.; Beek, van T.A.; Waard, de P.

    2005-01-01

    A rapid method for the simultaneous detection and identification of radical scavenging compounds in plant extracts was developed by combining an HPLC with on-line radical scavenging using DPPH as a model radical and an HPLC¿DAD¿SPE¿NMR system. Using this method a commercial rosemary extract was

  13. Rapid identification of Brucella isolates to the species level by real time PCR based single nucleotide polymorphism (SNP analysis

    Directory of Open Access Journals (Sweden)

    Smith Catherine J

    2008-06-01

    Full Text Available Abstract Background Brucellosis, caused by members of the genus Brucella, remains one of the world's major zoonotic diseases. Six species have classically been recognised within the family Brucella largely based on a combination of classical microbiology and host specificity, although more recently additional isolations of novel Brucella have been reported from various marine mammals and voles. Classical identification to species level is based on a biotyping approach that is lengthy, requires extensive and hazardous culturing and can be difficult to interpret. Here we describe a simple and rapid approach to identification of Brucella isolates to the species level based on real-time PCR analysis of species-specific single nucleotide polymorphisms (SNPs that were identified following a robust and extensive phylogenetic analysis of the genus. Results Seven pairs of short sequence Minor Groove Binding (MGB probes were designed corresponding to SNPs shown to possess an allele specific for each of the six classical Brucella spp and the marine mammal Brucella. Assays were optimised to identical reaction parameters in order to give a multiple outcome assay that can differentiate all the classical species and Brucella isolated from marine mammals. The scope of the assay was confirmed by testing of over 300 isolates of Brucella, all of which typed as predicted when compared to other phenotypic and genotypic approaches. The assay is sensitive being capable of detecting and differentiating down to 15 genome equivalents. We further describe the design and testing of assays based on three additional SNPs located within the 16S rRNA gene that ensure positive discrimination of Brucella from close phylogenetic relatives on the same platform. Conclusion The multiple-outcome assay described represents a new tool for the rapid, simple and unambiguous characterisation of Brucella to the species level. Furthermore, being based on a robust phylogenetic framework, the

  14. Investigation of Rapid Low-Power Microwave-Induction Heating Scheme on the Cross-Linking Process of the Poly(4-vinylphenol) for the Gate Insulator of Pentacene-Based Thin-Film Transistors.

    Science.gov (United States)

    Fan, Ching-Lin; Shang, Ming-Chi; Wang, Shea-Jue; Hsia, Mao-Yuan; Lee, Win-Der; Huang, Bohr-Ran

    2017-07-03

    In this study, a proposed Microwave-Induction Heating (MIH) scheme has been systematically studied to acquire suitable MIH parameters including chamber pressure, microwave power and heating time. The proposed MIH means that the thin indium tin oxide (ITO) metal below the Poly(4-vinylphenol) (PVP) film is heated rapidly by microwave irradiation and the heated ITO metal gate can heat the PVP gate insulator, resulting in PVP cross-linking. It is found that the attenuation of the microwave energy decreases with the decreasing chamber pressure. The optimal conditions are a power of 50 W, a heating time of 5 min, and a chamber pressure of 20 mTorr. When suitable MIH parameters were used, the effect of PVP cross-linking and the device performance were similar to those obtained using traditional oven heating, even though the cross-linking time was significantly decreased from 1 h to 5 min. Besides the gate leakage current, the interface trap state density (Nit) was also calculated to describe the interface status between the gate insulator and the active layer. The lowest interface trap state density can be found in the device with the PVP gate insulator cross-linked by using the optimal MIH condition. Therefore, it is believed that the MIH scheme is a good candidate to cross-link the PVP gate insulator for organic thin-film transistor applications as a result of its features of rapid heating (5 min) and low-power microwave-irradiation (50 W).

  15. Investigation of Rapid Low-Power Microwave-Induction Heating Scheme on the Cross-Linking Process of the Poly(4-vinylphenol for the Gate Insulator of Pentacene-Based Thin-Film Transistors

    Directory of Open Access Journals (Sweden)

    Ching-Lin Fan

    2017-07-01

    Full Text Available In this study, a proposed Microwave-Induction Heating (MIH scheme has been systematically studied to acquire suitable MIH parameters including chamber pressure, microwave power and heating time. The proposed MIH means that the thin indium tin oxide (ITO metal below the Poly(4-vinylphenol (PVP film is heated rapidly by microwave irradiation and the heated ITO metal gate can heat the PVP gate insulator, resulting in PVP cross-linking. It is found that the attenuation of the microwave energy decreases with the decreasing chamber pressure. The optimal conditions are a power of 50 W, a heating time of 5 min, and a chamber pressure of 20 mTorr. When suitable MIH parameters were used, the effect of PVP cross-linking and the device performance were similar to those obtained using traditional oven heating, even though the cross-linking time was significantly decreased from 1 h to 5 min. Besides the gate leakage current, the interface trap state density (Nit was also calculated to describe the interface status between the gate insulator and the active layer. The lowest interface trap state density can be found in the device with the PVP gate insulator cross-linked by using the optimal MIH condition. Therefore, it is believed that the MIH scheme is a good candidate to cross-link the PVP gate insulator for organic thin-film transistor applications as a result of its features of rapid heating (5 min and low-power microwave-irradiation (50 W.

  16. Rapid Detection and Identification of miRNAs by Surface-Enhanced Raman Spectroscopy Using Hollow Au Nanoflowers Substrates

    Directory of Open Access Journals (Sweden)

    Xiaowei Cao

    2017-01-01

    Full Text Available MicroRNAs (miRNAs are recognized as regulators of gene expression during the biological processes of cells as well as biomarkers of many diseases. Development of rapid and sensitive miRNA profiling methods is crucial for evaluating the pattern of miRNA expression related to normal and diseased states. This work presents a novel hollow Au nanoflowers (HAuNFs substrate for rapid detection and identification of miRNAs by surface-enhanced Raman scattering (SERS spectroscopy. We synthesized the HAuNFs by a seed-mediated growth approach. Then, HAuNFs substrates were fabricated by depositing HAuNFs onto the surfaces of (3-aminopropyltriethoxysilane- (APTES- functionalized ITO glass. The result demonstrated that HAuNFs substrates had very good reproducibility, homogeneous SERS activity, and high SERS effect. The substrates enabled us to successfully obtain the SERS spectra of miR-10a-5p, miR-125a-5p, and miR-196a-5p. The difference spectra among the three kinds of miRNAs were studied to better interpret the spectral differences and identify miRNA expression patterns with high accuracy. The principal component analysis (PCA of the SERS spectra was used to distinguish among the three kinds of miRNAs. Considering its time efficiency, being label-free, and its sensitivity, the SERS based on HAuNFs substrates is very promising for miRNA research and plays an important role in early disease detection and prevention.

  17. Saponin promotes rapid identification and antimicrobial susceptibility profiling of Gram-positive and Gram-negative bacteria in blood cultures with the Vitek 2 system.

    Science.gov (United States)

    Lupetti, A; Barnini, S; Morici, P; Ghelardi, E; Nibbering, P H; Campa, M

    2013-04-01

    The rapid identification and antimicrobial susceptibility testing (AST) of bacteria in clinical blood cultures is crucial to optimise antimicrobial therapy. A previous study involving small sample numbers revealed that the addition of saponin to blood cultures, further referred to as the new method, shortened considerably the turn-around time for the identification and AST of Gram-positive cocci as compared to the current method involving an overnight subculture. Here, we extend previous results and compare the identification and AST of blood cultures containing Gram-negative bacilli by the new and current methods. The identification and AST of 121 Gram-positive and 109 Gram-negative bacteria in clinical monomicrobial blood cultures by the new and current methods and, in the case of Gram-negative bacilli, by direct (no additions) inoculation into an automated system (rapid method) was assessed using the Vitek 2 system. Discrepancies between the results obtained with the different methods were solved by manual methods. The new method correctly identified 88 % of Gram-positive and 98 % of Gram-negative bacteria, and the rapid method correctly identified 94 % of Gram-negative bacteria. The AST for all antimicrobials by the new method were concordant with the current method for 55 % and correct for an additional 9 % of Gram-positive bacteria, and concordant with the current method for 62 % and correct for an additional 21 % of Gram-negative bacilli. The AST by the rapid method was concordant with the current method for 62 % and correct for an additional 12 % of Gram-negative bacilli. Together, saponin-treated monomicrobial blood cultures allow rapid and reliable identification and AST of Gram-positive and Gram-negative bacteria.

  18. Rapid identification of bacteria from positive blood culture bottles by MALDI-TOF MS following short-term incubation on solid media.

    Science.gov (United States)

    Altun, Osman; Botero-Kleiven, Silvia; Carlsson, Sarah; Ullberg, Måns; Özenci, Volkan

    2015-11-01

    Rapid identification of bacteria from blood cultures enables early initiation of appropriate antibiotic treatment in patients with bloodstream infections (BSI). The objective of the present study was to evaluate the use of matrix-associated laser desorption ionization-time of flight (MALDI-TOF) MS after a short incubation on solid media for rapid identification of bacteria from positive blood culture bottles. MALDI-TOF MS was performed after 2.5 and 5.5 h plate incubation of samples from positive blood cultures. Identification scores with values ≥ 1.7 were accepted as successful identification if the results were confirmed by conventional methods. Conventional methods included MALDI-TOF MS, Vitek 2, and diverse biochemical and agglutination tests after overnight culture. In total, 515 positive blood cultures with monomicrobial bacterial growth representing one blood culture per patient were included in the study. There were 229/515 (44.5%) and 286/515 (55.5%) blood culture bottles with Gram-negative bacteria (GNB) and Gram-positive bacteria (GPB), respectively. MALDI-TOF MS following short-term culture could accurately identify 300/515 (58.3%) isolates at 2.5 h, GNB being identified in greater proportion (180/229; 78.6%) than GPB (120/286; 42.0%). In an additional 124/515 bottles (24.1%), identification was successful at 5.5 h, leading to accurate identification of bacteria from 424/515 (82.3%) blood cultures after short-term culture. Interestingly, 11/24 of the isolated anaerobic bacteria could be identified after 5.5 h. The present study demonstrates, in a large number of clinical samples, that MALDI-TOF MS following short-term culture on solid medium is a reliable and rapid method for identification of bacteria from blood culture bottles with monomicrobial bacterial growth.

  19. Rapid identification of regulated organic chemical compounds in toys using ambient ionization and a miniature mass spectrometry system.

    Science.gov (United States)

    Guo, Xiangyu; Bai, Hua; Lv, Yueguang; Xi, Guangcheng; Li, Junfang; Ma, Xiaoxiao; Ren, Yue; Ouyang, Zheng; Ma, Qiang

    2018-04-01

    Rapid, on-site analysis was achieved through significantly simplified operation procedures for a wide variety of toy samples (crayon, temporary tattoo sticker, finger paint, modeling clay, and bubble solution) using a miniature mass spectrometry system with ambient ionization capability. The labor-intensive analytical protocols involving sample workup and chemical separation, traditionally required for MS-based analysis, were replaced by direct sampling analysis using ambient ionization methods. A Mini β ion trap miniature mass spectrometer was coupled with versatile ambient ionization methods, e.g. paper spray, extraction spray and slug-flow microextraction nanoESI for direct identification of prohibited colorants, carcinogenic primary aromatic amines, allergenic fragrances, preservatives and plasticizers from raw toy samples. The use of paper substrates coated with Co 3 O 4 nanoparticles allowed a great increase in sensitivity for paper spray. Limits of detection as low as 5μgkg -1 were obtained for target analytes. The methods being developed based on the integration of ambient ionization with miniature mass spectrometer represent alternatives to current in-lab MS analysis operation, and would enable fast, outside-the-lab screening of toy products to ensure children's safety and health. Copyright © 2017 Elsevier B.V. All rights reserved.

  20. Rapid Identification of Coumarins from Micromelum falcatum by UPLC-HRMS/MS and Targeted Isolation of Three New Derivatives

    Directory of Open Access Journals (Sweden)

    Eirini Kouloura

    2014-09-01

    Full Text Available Micromelum falcatum, a medicinal plant of the Rutaceae family, has been used in the Traditional Chinese Medicine (TCM mainly against colds and rheumatoid arthritis. Despite its traditional use the association of its constituents with possible anti-inflammatory activity has not been explored. During this study, a rapid UPLC-ESI(+-HRMS method was developed for the profiling of M. falcatum leave extracts and the targeted isolation of coumarin constituents. Based on chromatographic, spectroscopic and spectrometric features several 7-oxygenated coumarin derivatives were detected. After targeted isolation, eight coumarins, among them three new natural products, namely microfalcrin, microcoumaririn and micromelosidester, were purified using semi-preparative HPLC and unambiguously identified by 1 and 2D NMR. Furthermore, important spectrometric characteristics were revealed based on the HRMS and HRMS/MS spectra of the isolated 7-oxygenated coumarins facilitating their identification in complex mixtures. Finally, the anti-inflammatory properties of the extracts and representative compounds were evaluated by measuring the inhibition of the pro-inflammatory mediator NF-κB induction and nitric oxide (NO production.

  1. Rapid identification of coumarins from Micromelum falcatum by UPLC-HRMS/MS and targeted isolation of three new derivatives.

    Science.gov (United States)

    Kouloura, Eirini; Danika, Eirini; Kim, Sothea; Hoerlé, Mélanie; Cuendet, Muriel; Halabalaki, Maria; Skaltsounis, Leandros A

    2014-09-19

    Micromelum falcatum, a medicinal plant of the Rutaceae family, has been used in the Traditional Chinese Medicine (TCM) mainly against colds and rheumatoid arthritis. Despite its traditional use the association of its constituents with possible anti-inflammatory activity has not been explored. During this study, a rapid UPLC-ESI(+)-HRMS method was developed for the profiling of M. falcatum leave extracts and the targeted isolation of coumarin constituents. Based on chromatographic, spectroscopic and spectrometric features several 7-oxygenated coumarin derivatives were detected. After targeted isolation, eight coumarins, among them three new natural products, namely microfalcrin, microcoumaririn and micromelosidester, were purified using semi-preparative HPLC and unambiguously identified by 1 and 2D NMR. Furthermore, important spectrometric characteristics were revealed based on the HRMS and HRMS/MS spectra of the isolated 7-oxygenated coumarins facilitating their identification in complex mixtures. Finally, the anti-inflammatory properties of the extracts and representative compounds were evaluated by measuring the inhibition of the pro-inflammatory mediator NF-κB induction and nitric oxide (NO) production.

  2. Rapid Detection and Identification of Overdose Drugs in Saliva by Surface-Enhanced Raman Scattering Using Fused Gold Colloids

    Directory of Open Access Journals (Sweden)

    Frank Inscore

    2011-07-01

    Full Text Available The number of drug-related emergency room visits in the United States doubled from 2004 to 2009 to 4.6 million. Consequently there is a critical need to rapidly identify the offending drug(s, so that the appropriate medical care can be administered. In an effort to meet this need we have been investigating the ability of surface-enhanced Raman spectroscopy (SERS to detect and identify numerous drugs in saliva at ng/mL concentrations within 10 minutes. Identification is provided by matching measured spectra to a SERS library comprised of over 150 different drugs, each of which possess a unique spectrum. Trace detection is provided by fused gold colloids trapped within a porous glass matrix that generate SERS. Speed is provided by a syringe-driven sample system that uses a solid-phase extraction capillary combined with a SERS-active capillary in series. Spectral collection is provided by a portable Raman analyzer. Here we describe successful measurement of representative illicit, prescribed, and over-the-counter drugs by SERS, and 50 ng/mL cocaine in saliva as part of a focused study.

  3. Rapid detection and identification of 12 respiratory viruses using a dual priming oligonucleotide system-based multiplex PCR assay.

    Science.gov (United States)

    Kim, Suk Ran; Ki, Chang-Seok; Lee, Nam Yong

    2009-03-01

    Acute viral respiratory infections are among the most common causes of human disease. Rapid and accurate diagnosis of viral respiratory infections is important for providing timely therapeutic interventions. This study evaluated a new multiplex PCR assay (Seegene Inc., Seoul, Korea) for simultaneous detection and identification of 12 respiratory viruses using two primer mixes. The viruses included parainfluenza viruses 1, 2, and 3, human metapneumovirus, human coronavirus 229E/NL63 and OC43, adenovirus, influenza viruses A and B, human respiratory syncytial viruses A and B, and human rhinovirus A. The analytical sensitivity of the assay was 10-100 copies per reaction for each type of virus. There was no cross-reactivity with common bacterial or viral pathogens. A comparison with conventional viral culture and immunofluorescence was carried out using 101 respiratory specimens from 92 patients. Using viral culture, 57 specimens (56.4%) were positive without co-infection. The same viruses were identified in all 57 specimens using the multiplex PCR. Seven of the 57 specimens (12.3%) were found to be co-infected with other respiratory viruses, and 19 of 44 (43.2%) specimens which were negative by culture were positive by the multiplex PCR. The Seeplex Respiratory Virus Detection assay represents a significant improvement over the conventional methods for the detection of a broad spectrum of respiratory viruses.

  4. Rapid and inexpensive body fluid identification by RNA profiling-based multiplex High Resolution Melt (HRM) analysis

    National Research Council Canada - National Science Library

    Hanson, Erin K; Ballantyne, Jack

    2014-01-01

    .... However, traditional protein-based methods do not permit the identification of all body fluids and tissues, and thus molecular based strategies for the conclusive identification of all forensically...

  5. Rapid and inexpensive body fluid identification by RNA profiling-based multiplex High Resolution Melt (HRM) analysis

    National Research Council Canada - National Science Library

    Hanson, Erin K; Ballantyne, Jack

    2013-01-01

    .... However, traditional protein-based methods do not permit the identification of all body fluids and tissues, and thus molecular based strategies for the conclusive identification of all forensically...

  6. A simple method for rapid microbial identification from positive monomicrobial blood culture bottles through matrix-assisted laser desorption ionization time-of-flight mass spectrometry.

    Science.gov (United States)

    Lin, Jung-Fu; Ge, Mao-Cheng; Liu, Tsui-Ping; Chang, Shih-Cheng; Lu, Jang-Jih

    2017-06-30

    Rapid identification of microbes in the bloodstream is crucial in managing septicemia because of its high disease severity, and direct identification from positive blood culture bottles through matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) can shorten the turnaround time. Therefore, we developed a simple method for rapid microbiological identification from positive blood cultures by using MALDI-TOF MS. We modified previously developed methods to propose a faster, simpler and more economical method, which includes centrifugation and hemolysis. Specifically, our method comprises two-stage centrifugation with gravitational acceleration (g) at 600g and 3000g, followed by the addition of a lysis buffer and another 3000g centrifugation. In total, 324 monomicrobial bacterial cultures were identified. The success rate of species identification was 81.8%, which is comparable with other complex methods. The identification success rate was the highest for Gram-negative aerobes (85%), followed by Gram-positive aerobes (78.2%) and anaerobes (67%). The proposed method requires less than 10 min, costs less than US$0.2 per usage, and facilitates batch processing. We conclude that this method is feasible for clinical use in microbiology laboratories, and can serve as a reference for treatments or further complementary diagnostic testing. Copyright © 2017. Published by Elsevier B.V.

  7. A Rapid Identification Method for Calamine Using Near-Infrared Spectroscopy Based on Multi-Reference Correlation Coefficient Method and Back Propagation Artificial Neural Network.

    Science.gov (United States)

    Sun, Yangbo; Chen, Long; Huang, Bisheng; Chen, Keli

    2017-07-01

    As a mineral, the traditional Chinese medicine calamine has a similar shape to many other minerals. Investigations of commercially available calamine samples have shown that there are many fake and inferior calamine goods sold on the market. The conventional identification method for calamine is complicated, therefore as a result of the large scale of calamine samples, a rapid identification method is needed. To establish a qualitative model using near-infrared (NIR) spectroscopy for rapid identification of various calamine samples, large quantities of calamine samples including crude products, counterfeits and processed products were collected and correctly identified using the physicochemical and powder X-ray diffraction method. The NIR spectroscopy method was used to analyze these samples by combining the multi-reference correlation coefficient (MRCC) method and the error back propagation artificial neural network algorithm (BP-ANN), so as to realize the qualitative identification of calamine samples. The accuracy rate of the model based on NIR and MRCC methods was 85%; in addition, the model, which took comprehensive multiple factors into consideration, can be used to identify crude calamine products, its counterfeits and processed products. Furthermore, by in-putting the correlation coefficients of multiple references as the spectral feature data of samples into BP-ANN, a BP-ANN model of qualitative identification was established, of which the accuracy rate was increased to 95%. The MRCC method can be used as a NIR-based method in the process of BP-ANN modeling.

  8. Rapid Identification of Microorganisms from Positive Blood Culture by MALDI-TOF MS After Short-Term Incubation on Solid Medium.

    Science.gov (United States)

    Curtoni, Antonio; Cipriani, Raffaella; Marra, Elisa Simona; Barbui, Anna Maria; Cavallo, Rossana; Costa, Cristina

    2017-01-01

    Matrix-assisted laser-desorption/ionization time-of-flight (MALDI-TOF) mass spectrometry (MS) is a useful tool for rapid identification of microorganisms. Unfortunately, its direct application to positive blood culture is still lacking standardized procedures. In this study, we evaluated an easy- and rapid-to-perform protocol for MALDI-TOF MS direct identification of microorganisms from positive blood culture after a short-term incubation on solid medium. This protocol was used to evaluate direct identification of microorganisms from 162 positive monomicrobial blood cultures; at different incubation times (3, 5, 24 h), MALDI-TOF MS assay was performed from the growing microorganism patina. Overall, MALDI-TOF MS concordance with conventional methods at species level was 60.5, 80.2, and 93.8% at 3, 5, and 24 h, respectively. Considering only bacteria, the identification performances at species level were 64.1, 85.0, and 94.1% at 3, 5, and 24 h, respectively. This protocol applied to a commercially available MS typing system may represent, a fast and powerful diagnostic tool for pathogen direct identification and for a promptly and pathogen-driven antimicrobial therapy in selected cases.

  9. Rapid Identification of the Foodborne Pathogen Trichinella spp. by Matrix-Assisted Laser Desorption/Ionization Mass Spectrometry.

    Science.gov (United States)

    Mayer-Scholl, Anne; Murugaiyan, Jayaseelan; Neumann, Jennifer; Bahn, Peter; Reckinger, Sabine; Nöckler, Karsten

    2016-01-01

    Human trichinellosis occurs through consumption of raw or inadequately processed meat or meat products containing larvae of the parasitic nematodes of the genus Trichinella. Currently, nine species and three genotypes are recognized, of which T. spiralis, T. britovi and T. pseudospiralis have the highest public health relevance. To date, the differentiation of the larvae to the species and genotype level is based primarily on molecular methods, which can be relatively time consuming and labor intensive. Due to its rapidness and ease of use a matrix assisted laser desorption / ionization time of flight mass spectrometry (MALDI-TOF MS) reference spectra database using Trichinella strains of all known species and genotypes was created. A formicacid/acetonitrile protein extraction was carried out after pooling 10 larvae of each Trichinella species and genotype. Each sample was spotted 9 times using α-cyano 4-hydoxy cinnamic acid matrix and a MicroFlex LT mass spectrometer was used to acquire 3 spectra (m/z 2000 to 20000 Da) from each spot resulting in 27 spectra/species or genotype. Following the spectra quality assessment, Biotyper software was used to create a main spectra library (MSP) representing nine species and three genotypes of Trichinella. The evaluation of the spectra generated by MALDI-TOF MS revealed a classification which was comparable to the results obtained by molecular methods. Also, each Trichinella species utilized in this study was distinct and distinguishable with a high confidence level. Further, different conservation methods such as freezing and conservation in alcohol and the host species origin of the isolated larvae did not have a significant influence on the generated spectra. Therefore, the described MALDI-TOF MS can successfully be implemented for both genus and species level identification and represents a major step forward in the use of this technique in foodborne parasitology.

  10. Bioluminescence-based identification of nisin producers - a rapid and simple screening method for nisinogenic bacteria in food samples.

    Science.gov (United States)

    Virolainen, Nina; Guglielmetti, Simone; Arioli, Stefania; Karp, Matti

    2012-08-17

    We present a simple and rapid method for screening nisin producers that directly identifies nisinogenic bacteria by induction of bioluminescence within the Lactococcus lactis NZ9800lux biosensor strain (Immonen and Karp, 2007, Biosensors and Bioelectronics 22, 1982-7). An overlay of putative nisinogenic colonies with the biosensor strain gives identification results within 1h. Functionality and specificity of the method were verified by screening nisin producers among 144 raw milk colonies and a panel of 91 lactococcal strains. Studies performed on strains and colonies that did not induce bioluminescence but inhibited growth of the biosensor demonstrated that only nisinogenic bacteria can cause induction. Bacteria known to produce bacteriocins other than nisin failed to induce bioluminescence, further verifying the specificity of the assay. We discovered a non-inducing but inhibitory lactococcal strain harboring a modified nisin Z gene, and demonstrated that the source of the inhibitory action is not a non-inducing variant of nisin, but a bacteriocin of lower molecular weight. The concentration of nisin producers in a raw milk sample was 1.3 × 10(2)CFU/ml. We identified from raw milk a total of seven nisin Z producing L. lactis subsp. lactis colonies, which were shown by genetic fingerprinting to belong to three different groups. Among the panel of 91 lactococci, four strains were nisin A producers, and one strain harbored the modified nisin Z gene. The method presented here is robust, cost-effective and simple to perform, and avoids the pitfalls of traditional screening methods by directly specifying the identity of the inhibitory substance. Copyright © 2012 Elsevier B.V. All rights reserved.

  11. Rapid identification of drug-type strains in Cannabis sativa using loop-mediated isothermal amplification assay.

    Science.gov (United States)

    Kitamura, Masashi; Aragane, Masako; Nakamura, Kou; Watanabe, Kazuhito; Sasaki, Yohei

    2017-01-01

    In Cannabis sativa L., tetrahydrocannabinol (THC) is the primary psychoactive compound and exists as the carboxylated form, tetrahydrocannabinolic acid (THCA). C. sativa is divided into two strains based on THCA content-THCA-rich (drug-type) strains and THCA-poor (fiber-type) strains. Both strains are prohibited by law in many countries including Japan, whereas the drug-type strains are regulated in Canada and some European countries. As the two strains cannot be discriminated by morphological analysis, a simple method for identifying the drug-type strains is required for quality control in legal cultivation and forensic investigation. We have developed a novel loop-mediated isothermal amplification (LAMP) assay for identifying the drug-type strains of C. sativa. We designed two selective LAMP primer sets for on-site or laboratory use, which target the drug-type THCA synthase gene. The LAMP assay was accomplished within approximately 40 min. The assay showed high specificity for the drug-type strains and its sensitivity was the same as or higher than that of conventional polymerase chain reaction. We also showed the effectiveness of melting curve analysis that was conducted after the LAMP assay. The melting temperature values of the drug-type strains corresponded to those of the cloned drug-type THCA synthase gene, and were clearly different from those of the cloned fiber-type THCA synthase gene. Moreover, the LAMP assay with simple sample preparation could be accomplished within 1 h from sample treatment to identification without the need for special devices or techniques. Our rapid, sensitive, specific, and simple assay is expected to be applicable to laboratory and on-site detection.

  12. Rapid Identification of the Foodborne Pathogen Trichinella spp. by Matrix-Assisted Laser Desorption/Ionization Mass Spectrometry.

    Directory of Open Access Journals (Sweden)

    Anne Mayer-Scholl

    Full Text Available Human trichinellosis occurs through consumption of raw or inadequately processed meat or meat products containing larvae of the parasitic nematodes of the genus Trichinella. Currently, nine species and three genotypes are recognized, of which T. spiralis, T. britovi and T. pseudospiralis have the highest public health relevance. To date, the differentiation of the larvae to the species and genotype level is based primarily on molecular methods, which can be relatively time consuming and labor intensive. Due to its rapidness and ease of use a matrix assisted laser desorption / ionization time of flight mass spectrometry (MALDI-TOF MS reference spectra database using Trichinella strains of all known species and genotypes was created. A formicacid/acetonitrile protein extraction was carried out after pooling 10 larvae of each Trichinella species and genotype. Each sample was spotted 9 times using α-cyano 4-hydoxy cinnamic acid matrix and a MicroFlex LT mass spectrometer was used to acquire 3 spectra (m/z 2000 to 20000 Da from each spot resulting in 27 spectra/species or genotype. Following the spectra quality assessment, Biotyper software was used to create a main spectra library (MSP representing nine species and three genotypes of Trichinella. The evaluation of the spectra generated by MALDI-TOF MS revealed a classification which was comparable to the results obtained by molecular methods. Also, each Trichinella species utilized in this study was distinct and distinguishable with a high confidence level. Further, different conservation methods such as freezing and conservation in alcohol and the host species origin of the isolated larvae did not have a significant influence on the generated spectra. Therefore, the described MALDI-TOF MS can successfully be implemented for both genus and species level identification and represents a major step forward in the use of this technique in foodborne parasitology.

  13. Rapid identification of cell-specific, internalizing RNA aptamers with bioinformatics analyses of a cell-based aptamer selection.

    Directory of Open Access Journals (Sweden)

    William H Thiel

    Full Text Available The broad applicability of RNA aptamers as cell-specific delivery tools for therapeutic reagents depends on the ability to identify aptamer sequences that selectively access the cytoplasm of distinct cell types. Towards this end, we have developed a novel approach that combines a cell-based selection method (cell-internalization SELEX with high-throughput sequencing (HTS and bioinformatics analyses to rapidly identify cell-specific, internalization-competent RNA aptamers.We demonstrate the utility of this approach by enriching for RNA aptamers capable of selective internalization into vascular smooth muscle cells (VSMCs. Several rounds of positive (VSMCs and negative (endothelial cells; ECs selection were performed to enrich for aptamer sequences that preferentially internalize into VSMCs. To identify candidate RNA aptamer sequences, HTS data from each round of selection were analyzed using bioinformatics methods: (1 metrics of selection enrichment; and (2 pairwise comparisons of sequence and structural similarity, termed edit and tree distance, respectively. Correlation analyses of experimentally validated aptamers or rounds revealed that the best cell-specific, internalizing aptamers are enriched as a result of the negative selection step performed against ECs.We describe a novel approach that combines cell-internalization SELEX with HTS and bioinformatics analysis to identify cell-specific, cell-internalizing RNA aptamers. Our data highlight the importance of performing a pre-clear step against a non-target cell in order to select for cell-specific aptamers. We expect the extended use of this approach to enable the identification of aptamers to a multitude of different cell types, thereby facilitating the broad development of targeted cell therapies.

  14. Rapid identification of cell-specific, internalizing RNA aptamers with bioinformatics analyses of a cell-based aptamer selection.

    Science.gov (United States)

    Thiel, William H; Bair, Thomas; Peek, Andrew S; Liu, Xiuying; Dassie, Justin; Stockdale, Katie R; Behlke, Mark A; Miller, Francis J; Giangrande, Paloma H

    2012-01-01

    The broad applicability of RNA aptamers as cell-specific delivery tools for therapeutic reagents depends on the ability to identify aptamer sequences that selectively access the cytoplasm of distinct cell types. Towards this end, we have developed a novel approach that combines a cell-based selection method (cell-internalization SELEX) with high-throughput sequencing (HTS) and bioinformatics analyses to rapidly identify cell-specific, internalization-competent RNA aptamers. We demonstrate the utility of this approach by enriching for RNA aptamers capable of selective internalization into vascular smooth muscle cells (VSMCs). Several rounds of positive (VSMCs) and negative (endothelial cells; ECs) selection were performed to enrich for aptamer sequences that preferentially internalize into VSMCs. To identify candidate RNA aptamer sequences, HTS data from each round of selection were analyzed using bioinformatics methods: (1) metrics of selection enrichment; and (2) pairwise comparisons of sequence and structural similarity, termed edit and tree distance, respectively. Correlation analyses of experimentally validated aptamers or rounds revealed that the best cell-specific, internalizing aptamers are enriched as a result of the negative selection step performed against ECs. We describe a novel approach that combines cell-internalization SELEX with HTS and bioinformatics analysis to identify cell-specific, cell-internalizing RNA aptamers. Our data highlight the importance of performing a pre-clear step against a non-target cell in order to select for cell-specific aptamers. We expect the extended use of this approach to enable the identification of aptamers to a multitude of different cell types, thereby facilitating the broad development of targeted cell therapies.

  15. DNA microarray-based solid-phase RT-PCR for rapid detection and identification of influenza virus type A and subtypes H5 and H7

    DEFF Research Database (Denmark)

    Yi, Sun; Dhumpa, Raghuram; Bang, Dang Duong

    2011-01-01

    Endemic of avian influenza virus (AIV) in Asia and epizootics in some European regions have caused considerable public concern on a possible pandemic of AIV. A rapid method for virus detection and effective surveillance in wild avian, poultry production as well as in humans is required....... In this article, a DNA microarray-based solid-phase polymerase chain reaction (PCR) approach has been developed for rapid detection of influenza virus type A and for simultaneous identification of pathogenic virus subtypes H5 and H7. This solid-phase RT-PCR method combined reverse-transcription amplification...

  16. Novel toggle-rate based energy-efficient scheme for heavy load real-time IM-DD OFDM-PON with ONU LLID identification in time-domain using amplitude decision.

    Science.gov (United States)

    Qin, Youxiang; Zhang, Junjie

    2017-07-10

    A novel low complexity and energy-efficient scheme by controlling the toggle-rate of ONU with time-domain amplitude identification is proposed for a heavy load downlink in an intensity-modulation and direct-detection orthogonal frequency division multiplexing passive optical network (IM-DD OFDM-PON). In a conventional OFDM-PON downlink, all ONUs have to perform demodulation for all the OFDM frames in a broadcast way no matter whether the frames are targeted to or not, which causes a huge energy waste. However, in our scheme, the optical network unit (ONU) logical link identifications (LLIDs) are inserted into each downlink OFDM frame in time-domain at the optical line terminal (OLT) side. At the ONU side, the LLID is obtained with a low complexity and high precision amplitude identification method. The ONU sets the toggle-rate of demodulation module to zero when the frames are not targeted to, which avoids unnecessary digital signal processing (DSP) energy consumption. Compared with the sleep-mode methods consisting of clock recovery and synchronization, toggle-rate shows its advantage in fast changing, which is more suitable for the heavy load scenarios. Moreover, for the first time to our knowledge, the characteristics of the proposed scheme are investigated in a real-time IM-DD OFDM system, which performs well at the received optical power as low as -21dBm. The experimental results show that 25.1% energy consumption can be saved in the receiver compared to the conventional configurations.

  17. Rapid and inexpensive body fluid identification by RNA profiling-based multiplex High Resolution Melt (HRM) analysis

    OpenAIRE

    Erin K. Hanson; Jack Ballantyne

    2014-01-01

    Positive identification of the nature of biological material present on evidentiary items can be crucial for understanding the circumstances surrounding a crime. However, traditional protein-based methods do not permit the identification of all body fluids and tissues, and thus molecular based strategies for the conclusive identification of all forensically relevant biological fluids and tissues need to be developed. Messenger RNA (mRNA) profiling is an example of such a molecular-based appro...

  18. Rapid and accurate identification of Mycobacterium tuberculosis complex and common non-tuberculous mycobacteria by multiplex real-time PCR targeting different housekeeping genes.

    Science.gov (United States)

    Nasr Esfahani, Bahram; Rezaei Yazdi, Hadi; Moghim, Sharareh; Ghasemian Safaei, Hajieh; Zarkesh Esfahani, Hamid

    2012-11-01

    Rapid and accurate identification of mycobacteria isolates from primary culture is important due to timely and appropriate antibiotic therapy. Conventional methods for identification of Mycobacterium species based on biochemical tests needs several weeks and may remain inconclusive. In this study, a novel multiplex real-time PCR was developed for rapid identification of Mycobacterium genus, Mycobacterium tuberculosis complex (MTC) and the most common non-tuberculosis mycobacteria species including M. abscessus, M. fortuitum, M. avium complex, M. kansasii, and the M. gordonae in three reaction tubes but under same PCR condition. Genetic targets for primer designing included the 16S rDNA gene, the dnaJ gene, the gyrB gene and internal transcribed spacer (ITS). Multiplex real-time PCR was setup with reference Mycobacterium strains and was subsequently tested with 66 clinical isolates. Results of multiplex real-time PCR were analyzed with melting curves and melting temperature (T (m)) of Mycobacterium genus, MTC, and each of non-tuberculosis Mycobacterium species were determined. Multiplex real-time PCR results were compared with amplification and sequencing of 16S-23S rDNA ITS for identification of Mycobacterium species. Sensitivity and specificity of designed primers were each 100 % for MTC, M. abscessus, M. fortuitum, M. avium complex, M. kansasii, and M. gordonae. Sensitivity and specificity of designed primer for genus Mycobacterium was 96 and 100 %, respectively. According to the obtained results, we conclude that this multiplex real-time PCR with melting curve analysis and these novel primers can be used for rapid and accurate identification of genus Mycobacterium, MTC, and the most common non-tuberculosis Mycobacterium species.

  19. Rapid identification of Burkholderia pseudomallei and Burkholderia mallei by fluorescence in situ hybridization (FISH) from culture and paraffin-embedded tissue samples.

    Science.gov (United States)

    Hagen, Ralf M; Frickmann, Hagen; Elschner, Mandy; Melzer, Falk; Neubauer, Heinrich; Gauthier, Yves P; Racz, Paul; Poppert, Sven

    2011-11-01

    We evaluated newly developed probes for rapid identification of Burkholderia (B.) pseudomallei and B. mallei and differentiation from B. thailandensis by fluorescence in situ hybridization (FISH). FISH correctly identified 100% of the tested B. pseudomallei (11), B. mallei (11), and B. thailandensis (1) strains, excluded 100% of all tested negative controls (61), and allowed demonstration of B. pseudomallei infection in a paraffin-embedded spleen tissue sample of an experimentally infected mouse. Copyright © 2011 Elsevier GmbH. All rights reserved.

  20. Rapid identification of Candida spp. frequently involved in invasive mycoses by using flow-through hybridization and Gene Chip (FHGC) technology.

    Science.gov (United States)

    Li, Chen; Ding, Xiurong; Liu, Zhizhong; Zhu, Juanjuan

    2017-01-01

    The incidence of invasive fungal infections in immunocompromised patients has increased in recent decades. Rapid and accurate identification of these pathogenic fungi is crucial for initiating a timely, safe, and effective antifungal therapy. Here we developed a microarray based on flow-through hybridization gene chip technology. The microarray was tested for its specificity using a panel of reference and blinded clinical isolates. The results proved that this microarray was highly discriminative, leading to the unequivocal identification of each species, including Candida famata and the highly related species Candida parapsilosis, Candida orthopsilosis, and Candida metapsilosis. This new system represents a reliable method that is of potential use in clinical laboratories for the simultaneous detection and identification of the most common pathogenic fungi. Copyright © 2016 Elsevier B.V. All rights reserved.

  1. Rapid identification of Enterobacteriaceae in milk and dairy products with the Matrix-Assisted Laser Desorption Ionization–Time of Flight Mass Spectrometry (MALDI-TOF MS

    Directory of Open Access Journals (Sweden)

    Miroslava Kačániová

    2017-11-01

    Full Text Available Identification of microorganisms by MALDI-TOF MS Biotyper has been demonstrated to be accurate, rapid and lower cost than conventional food investigation methods. Rapid identification of pathogenic and spoilage microorganisms is crucial for dairy industry to ensure the quality and safety of milk and milk products. In this study, the bacterial species representing the Enterobacteriaceae family were identified in raw milk and milk products using the MALDI-TOF MS mass spectrometry. Altogether, 20 samples of Slovak milk and milk products were examined. Samples were cultured on VRBG agar at 37 °C for 24-48 h. Typical bacterial colonies were selected for identification with MALDI-TOF MS Biotyper. Escherichia coli and Enterobacter sp. were the most abundant  Enterobacteriaceae family representatives identified. E. coli was found in nine  and Enterobacter sp. in eight samples. Enterobacter sp. comprised 49 % and Escherichia coli 23 % of all bacterial isolates. The study shows that MALDI-TOF MS Biotyper was reliable identification method of Enterobacteriaceae in milk and dairy products. This method was helpful in evaluation of bacterial contamination of milk.

  2. Rapid Detection ofStaphylococcus aureusand Related Species Isolated from Food, Environment, Cosmetics, a Medical Device, and Clinical Samples Using the VITEK MS Microbial Identification System.

    Science.gov (United States)

    Sulaiman, Irshad M; Banerjee, Pratik; Hsieh, Ying-Hsin; Miranda, Nancy; Simpson, Steven; Kerdahi, Khalil

    2017-09-15

    Staphylococcus spp. is considered as one of the most common human-pathogenic bacteria, causing illnesses ranging from nonthreatening skin infections to lethal diseases, including sepsis, pneumonia, bloodstream infections, and food poisoning. The emergence of methicillin-resistant Staphylococcus aureus strains has increased morbidity and mortality and resulted in a major healthcare burden worldwide. Single and multilocus sequence typing have been extensively used in the identification of Staphylococcus species. Nevertheless, these assays are relatively time-consuming and require high-quality DNA. Matrix-assisted laser desorption ionization-time-of-flight has been used recently for the rapid identification of several bacterial species. In this study, we have examined 47 Staphylococcus isolates recovered from food, environment, clinical samples, cosmetic products, and a medical device and 3 American Type Culture Collection Staphylococcus reference isolates using bioMérieux VITEK MS and VITEK 2 systems to determine isolate identity. Sequencing of the 16S ribosomal RNA gene was performed to confirm and compare the species identification data generated by VITEK 2 and VITEK MS systems. Although the VITEK 2 system could not identify one of the isolates, VITEK MS identified all 50 Staphylococcus spp. isolates tested. Results of this study clearly suggest that VITEK MS can be used in the rapid identification of Staphylococcus isolates of public health importance.

  3. Evaluation of the BD MGIT TBc Identification Test (TBc ID), a rapid chromatographic immunoassay for the detection of Mycobacterium tuberculosis complex from liquid culture.

    Science.gov (United States)

    Martin, Anandi; Bombeeck, Deirdre; Fissette, Krista; de Rijk, Pim; Hernández-Neuta, Ivan; Del Portillo, Patricia; Palomino, Juan Carlos

    2011-02-01

    The BACTEC MGIT 960 system is increasingly used to culture Mycobacterium tuberculosis. We evaluated the performance of the new immunochromatographic assay BD MGIT TBc Identification Test (TBc ID) for the rapid identification of M. tuberculosis complex in clinical samples when performed directly from BACTEC MGIT 960 culture positive for acid-fast bacilli (AFB). Of 92 cultures evaluated, the sensitivity and specificity of the TBc ID test was 98.5% and 100%, respectively compared to sequencing of the 16S rRNA gene. One culture that was TBc ID test negative but that was identified as M. tuberculosis by 16S rRNA sequencing was confirmed to have a mutation in the mpt64 gene. The TBc ID test is an easy and sensitive method for the identification of M. tuberculosis complex in liquid culture medium, does not require a high level of skills, neither any additional specific equipment and gives results in 15 min, which provide a good alternative for the rapid identification of M. tuberculosis complex in liquid medium. Copyright © 2010 Elsevier B.V. All rights reserved.

  4. Rapid identification of clinical mycobacterial isolates by protein profiling using matrix assisted laser desorption ionization-time of flight mass spectrometry.

    Science.gov (United States)

    Panda, A; Kurapati, S; Samantaray, J C; Myneedu, V P; Verma, A; Srinivasan, A; Ahmad, H; Behera, D; Singh, U B

    2013-01-01

    The purpose of this study was to evaluate the identification of Mycobacterium tuberculosis which is often plagued with ambiguity. It is a time consuming process requiring 4-8 weeks after culture positivity, thereby delaying therapeutic intervention. For a successful treatment and disease management, timely diagnosis is imperative. We evaluated a rapid, proteomic based technique for identification of clinical mycobacterial isolates by protein profiling using matrix assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS). Freshly grown mycobacterial isolates were used. Acetonitrile/trifluoroacetic acid extraction procedure was carried out, following which cinnamic acid charged plates were subjected to identification by MALDI-TOF MS. A comparative analysis of 42 clinical mycobacterial isolates using the MALDI-TOF MS and conventional techniques was carried out. Among these, 97.61% were found to corroborate with the standard methods at genus level and 85.36% were accurate till the species level. One out of 42 was not in accord with the conventional assays because MALDI-TOF MS established it as Mycobacterium tuberculosis (log (score)>2.0) and conventional methods established it to be non-tuberculous Mycobacterium. MALDI-TOF MS was found to be an accurate, rapid, cost effective and robust system for identification of mycobacterial species. This innovative approach holds promise for early therapeutic intervention leading to better patient care.

  5. Rapid identification of Iranian Acinetobacter baumannii strains by single PCR assay using BLA oxa-51 -like carbapenemase and evaluation of the antimicrobial resistance profiles of the isolates.

    Science.gov (United States)

    Akbari, Mahdi; Mahdi, Akbari; Niakan, Mohammad; Mohammad, Niakan; Taherikalani, Morovat; Morovat, Taherikalani; Feizabadi, Mohammad Mehdi; Mhammad-Mahdi, Feizabadi; Azadi, Namam Ali; Namam-Ali, Azadi; Soroush, Setareh; Setareh, Soroush; Emaneini, Mohammad; Mohammad, Emaneini; Abdolkarimi, Amir; Amir, Abdolkarimi; Maleki, Abbas; Abbas, Maleki; Hematian, Ali; Ali, Hematian

    2010-06-01

    The rapid identification of relevant bacterial pathogens is of utmost importance in clinical settings. The aim of this study was to test a rapid identification technique for A. baumannii strains from Tehran Hospitals and to determine the antibiotic resistance profiles of the isolates. A hundred strains of Acinetobacter spp. grown from clinical specimens were identified as A. baumannii by conventional methods. Using PCR a bla OXA-51 -like gene was detected in all A. baumannii isolates but not in other species of acinetobacter. More than half of the isolates proved resistant to a variety of antibiotics by the disk diffusion technique. The rate of resistance to gentamicin, imipenem, ampicillin-sulbactam and amikacin was determined to be 45%, 53%, 62% and 62%, respectively. Moreover, most isolates (more than 90%) showed resistance to cephalosporins. This study shows that the demonstration of the bla OXA-51-like gene is a reliable and rapid way for the presumptive identification of A. baumannii and reveals that the rate of antibiotic resistance is high in Iranian A. baumannii isolates to a variety of antibiotics.

  6. [Rapid identification of Riemerella anatipestifer on the basis of specific PCR amplifying 16S rDNA].

    Science.gov (United States)

    Qu, Feng-fa; Cai, Chang; Zheng, Xian-jin; Zhang, Da-bing

    2006-02-01

    which were probably infected with RA were chosen to identify the accuracy and sensitivity of this PCR method. Several other conventional detecting methods including bacteria isolating, differentiating culture, biochemical experiments and serotyping were used at the same time. The templates of PCR were extracted from brains or livers by boiling method with Chelex 100. Finally, 3 cases were identified as RA infection by the conventional methods and 4 by the PCR method, which proved the good accuracy and sensitivity of the PCR method. Thus, this PCR assay provides a rapid and accurate method for identification of Riemerella anatipestifer. It will help to make the final decision in clinical diagnose or species identification, especially when a new serotype or sub-serotype of RA comes up.

  7. Efficient and privacy-preserving biometric identification in cloud

    Directory of Open Access Journals (Sweden)

    Changhee Hahn

    2016-09-01

    Full Text Available With the rapid growth in the development of smart devices equipped with biometric sensors, client identification system using biometric traits are widely adopted across various applications. Among many biometric traits, fingerprint-based identification systems have been extensively studied and deployed. However, to adopt biometric identification systems in practical applications, two main obstacles in terms of efficiency and client privacy must be resolved simultaneously. That is, identification should be performed at an acceptable time, and only a client should have access to his/her biometric traits, which are not revocable if leaked. Until now, multiple studies have demonstrated successful protection of client biometric data; however, such systems lack efficiency that leads to excessive time utilization for identification. The most recently researched scheme shows efficiency improvements but reveals client biometric traits to other entities such as biometric database server. This violates client privacy. In this paper, we propose an efficient and privacy-preserving fingerprint identification scheme by using cloud systems. The proposed scheme extensively exploits the computation power of a cloud so that most of the laborious computations are performed by the cloud service provider. According to our experimental results on an Amazon EC2 cloud, the proposed scheme is faster than the existing schemes and guarantees client privacy by exploiting symmetric homomorphic encryption. Our security analysis shows that during identification, the client fingerprint data is not disclosed to the cloud service provider or fingerprint database server.

  8. Matrix-assisted laser desorption/ionisation-time of flight mass spectrometry: rapid identification of bacteria isolated from patients with cystic fibrosis.

    Science.gov (United States)

    Baillie, S; Ireland, K; Warwick, S; Wareham, D; Wilks, M

    2013-01-01

    Despite extensive research into the diagnosis and management of cystic fibrosis (CF) over the past decades, sufferers still have a median life expectancy of less than 37 years. Respiratory tract infections have a significant role in increasing the morbidity and mortality of patients with CF via a progressive decline in lung function. Rapid identification of organisms recovered from CF sputum is necessary for effective management of respiratory tract infections; however, standard techniques of identification are slow, technically demanding and expensive. The aim of this study is to asses the suitability of matrix-assisted laser desorption/ionisation-time of flight mass spectrometry (MALDI-TOF MS) in identifying bacteria isolated from the respiratory tract of patients with CF, and is assessed by testing the accuracy of MALDI-TOF MS in identifying samples from a reference collection of rare CF strains in conjunction with comparing MALDI-TOF MS and standard techniques in identifying clinical isolates from sputum samples of CF patients. MALDI-TOF MS accurately identified 100% of isolates from the reference collection of rare CF pathogens (EuroCare CF collection). The isolate identification given by MALDI-TOF MS agreed with that given by standard techniques for 479/481 (99.6%) clinical isolates obtained from respiratory samples provided by patients with CE In two (0.4%) of 481 samples there was a discrepancy in identification between MALDI-TOF MS and standard techniques. One organism was identified as Pseudomonas aeruginosa by MALDI-TOF but could only be identified by the laboratory's standard methods as of the Pseudomonas genus. The second organism was identified as P. beteli by MALDI-TOF MS and Stenotrophomonas maltophilia by standard methods. This study shows that MALDI-TOF MS is superior to standard techniques in providing cheap, rapid and accurate identification of CF sputum isolates.

  9. Rapid identification of Burkholderia mallei and Burkholderia pseudomallei by intact cell Matrix-assisted Laser Desorption/Ionisation mass spectrometric typing

    Science.gov (United States)

    2012-01-01

    Background Burkholderia (B.) pseudomallei and B. mallei are genetically closely related species. B. pseudomallei causes melioidosis in humans and animals, whereas B. mallei is the causative agent of glanders in equines and rarely also in humans. Both agents have been classified by the CDC as priority category B biological agents. Rapid identification is crucial, because both agents are intrinsically resistant to many antibiotics. Matrix-assisted laser desorption/ionisation mass spectrometry (MALDI-TOF MS) has the potential of rapid and reliable identification of pathogens, but is limited by the availability of a database containing validated reference spectra. The aim of this study was to evaluate the use of MALDI-TOF MS for the rapid and reliable identification and differentiation of B. pseudomallei and B. mallei and to build up a reliable reference database for both organisms. Results A collection of ten B. pseudomallei and seventeen B. mallei strains was used to generate a library of reference spectra. Samples of both species could be identified by MALDI-TOF MS, if a dedicated subset of the reference spectra library was used. In comparison with samples representing B. mallei, higher genetic diversity among B. pseudomallei was reflected in the higher average Eucledian distances between the mass spectra and a broader range of identification score values obtained with commercial software for the identification of microorganisms. The type strain of B. pseudomallei (ATCC 23343) was isolated decades ago and is outstanding in the spectrum-based dendrograms probably due to massive methylations as indicated by two intensive series of mass increments of 14 Da specifically and reproducibly found in the spectra of this strain. Conclusions Handling of pathogens under BSL 3 conditions is dangerous and cumbersome but can be minimized by inactivation of bacteria with ethanol, subsequent protein extraction under BSL 1 conditions and MALDI-TOF MS analysis being faster than

  10. Rapid identification of Burkholderia mallei and Burkholderia pseudomallei by intact cell Matrix-assisted Laser Desorption/Ionisation mass spectrometric typing.

    Science.gov (United States)

    Karger, Axel; Stock, Rüdiger; Ziller, Mario; Elschner, Mandy C; Bettin, Barbara; Melzer, Falk; Maier, Thomas; Kostrzewa, Markus; Scholz, Holger C; Neubauer, Heinrich; Tomaso, Herbert

    2012-10-10

    Burkholderia (B.) pseudomallei and B. mallei are genetically closely related species. B. pseudomallei causes melioidosis in humans and animals, whereas B. mallei is the causative agent of glanders in equines and rarely also in humans. Both agents have been classified by the CDC as priority category B biological agents. Rapid identification is crucial, because both agents are intrinsically resistant to many antibiotics. Matrix-assisted laser desorption/ionisation mass spectrometry (MALDI-TOF MS) has the potential of rapid and reliable identification of pathogens, but is limited by the availability of a database containing validated reference spectra. The aim of this study was to evaluate the use of MALDI-TOF MS for the rapid and reliable identification and differentiation of B. pseudomallei and B. mallei and to build up a reliable reference database for both organisms. A collection of ten B. pseudomallei and seventeen B. mallei strains was used to generate a library of reference spectra. Samples of both species could be identified by MALDI-TOF MS, if a dedicated subset of the reference spectra library was used. In comparison with samples representing B. mallei, higher genetic diversity among B. pseudomallei was reflected in the higher average Eucledian distances between the mass spectra and a broader range of identification score values obtained with commercial software for the identification of microorganisms. The type strain of B. pseudomallei (ATCC 23343) was isolated decades ago and is outstanding in the spectrum-based dendrograms probably due to massive methylations as indicated by two intensive series of mass increments of 14 Da specifically and reproducibly found in the spectra of this strain. Handling of pathogens under BSL 3 conditions is dangerous and cumbersome but can be minimized by inactivation of bacteria with ethanol, subsequent protein extraction under BSL 1 conditions and MALDI-TOF MS analysis being faster than nucleic amplification methods. Our

  11. Rapid identification of Burkholderia mallei and Burkholderia pseudomallei by intact cell Matrix-assisted Laser Desorption/Ionisation mass spectrometric typing

    Directory of Open Access Journals (Sweden)

    Karger Axel

    2012-10-01

    Full Text Available Abstract Background Burkholderia (B. pseudomallei and B. mallei are genetically closely related species. B. pseudomallei causes melioidosis in humans and animals, whereas B. mallei is the causative agent of glanders in equines and rarely also in humans. Both agents have been classified by the CDC as priority category B biological agents. Rapid identification is crucial, because both agents are intrinsically resistant to many antibiotics. Matrix-assisted laser desorption/ionisation mass spectrometry (MALDI-TOF MS has the potential of rapid and reliable identification of pathogens, but is limited by the availability of a database containing validated reference spectra. The aim of this study was to evaluate the use of MALDI-TOF MS for the rapid and reliable identification and differentiation of B. pseudomallei and B. mallei and to build up a reliable reference database for both organisms. Results A collection of ten B. pseudomallei and seventeen B. mallei strains was used to generate a library of reference spectra. Samples of both species could be identified by MALDI-TOF MS, if a dedicated subset of the reference spectra library was used. In comparison with samples representing B. mallei, higher genetic diversity among B. pseudomallei was reflected in the higher average Eucledian distances between the mass spectra and a broader range of identification score values obtained with commercial software for the identification of microorganisms. The type strain of B. pseudomallei (ATCC 23343 was isolated decades ago and is outstanding in the spectrum-based dendrograms probably due to massive methylations as indicated by two intensive series of mass increments of 14 Da specifically and reproducibly found in the spectra of this strain. Conclusions Handling of pathogens under BSL 3 conditions is dangerous and cumbersome but can be minimized by inactivation of bacteria with ethanol, subsequent protein extraction under BSL 1 conditions and MALDI-TOF MS

  12. DNA-based identification of invasive alien species in relation to Canadian federal policy and law, and the basis of rapid-response management.

    Science.gov (United States)

    Thomas, Vernon G; Hanner, Robert H; Borisenko, Alex V

    2016-11-01

    Managing invasive alien species in Canada requires reliable taxonomic identification as the basis of rapid-response management. This can be challenging, especially when organisms are small and lack morphological diagnostic features. DNA-based techniques, such as DNA barcoding, offer a reliable, rapid, and inexpensive toolkit for taxonomic identification of individual or bulk samples, forensic remains, and even environmental DNA. Well suited for this requirement, they could be more broadly deployed and incorporated into the operating policy and practices of Canadian federal departments and should be authorized under these agencies' articles of law. These include Fisheries and Oceans Canada, Canadian Food Inspection Agency, Transport Canada, Environment Canada, Parks Canada, and Health Canada. These efforts should be harmonized with the appropriate provisions of provincial jurisdictions, for example, the Ontario Invasive Species Act. This approach necessitates that a network of accredited, certified laboratories exists, and that updated DNA reference libraries are readily accessible. Harmonizing this approach is vital among Canadian federal agencies, and between the federal and provincial levels of government. Canadian policy and law must also be harmonized with that of the USA when detecting, and responding to, invasive species in contiguous lands and waters. Creating capacity in legislation for use of DNA-based identifications brings the authority to fund, train, deploy, and certify staff, and to refine further developments in this molecular technology.

  13. DNA barcoding and morphological analysis for rapid identification of most economically important crop-infesting Sunn pests belonging to Eurygaster Laporte, 1833 (Hemiptera, Scutelleridae

    Directory of Open Access Journals (Sweden)

    Mikhail Y. Syromyatnikov

    2017-10-01

    Full Text Available The genus Eurygaster Laporte, 1833 includes ten species five of which inhabit the European part of Russia. The harmful species of the genus is E. integriceps. Eurygaster species identification based on the morphological traits is very difficult, while that of the species at the egg or larval stages is extremely difficult or impossible. Eurygaster integriceps, E. maura, and E. testudinaria differ only slightly between each other morphologically, E. maura and E. testudinaria being almost indiscernible. DNA barcoding based on COI sequences have shown that E. integriceps differs significantly from these closely related species, which enables its rapid and accurate identification. Based on COI nucleotide sequences, three species of Sunn pests, E. maura, E. testudinarius, E. dilaticollis, could not be differentiated from each other through DNA barcoding. The difference in the DNA sequences between the COI gene of E. integriceps and COI genes of E. maura and E. testudinarius was more than 4%. In the present study DNA barcoding of two Eurygaster species was performed for the first time on E. integriceps, the most dangerous pest in the genus, and E. dilaticollis that only inhabits natural ecosystems. The PCR-RFLP method was developed in this work for the rapid identification of E. integriceps.

  14. DNA barcoding and morphological analysis for rapid identification of most economically important crop-infesting Sunn pests belonging to Eurygaster Laporte, 1833 (Hemiptera, Scutelleridae).

    Science.gov (United States)

    Syromyatnikov, Mikhail Y; Golub, Victor B; Kokina, Anastasia V; Victoria A Soboleva; Popov, Vasily N

    2017-01-01

    The genus Eurygaster Laporte, 1833 includes ten species five of which inhabit the European part of Russia. The harmful species of the genus is E. integriceps. Eurygaster species identification based on the morphological traits is very difficult, while that of the species at the egg or larval stages is extremely difficult or impossible. Eurygaster integriceps, E. maura, and E. testudinaria differ only slightly between each other morphologically, E. maura and E. testudinaria being almost indiscernible. DNA barcoding based on COI sequences have shown that E. integriceps differs significantly from these closely related species, which enables its rapid and accurate identification. Based on COI nucleotide sequences, three species of Sunn pests, E. maura, E. testudinarius, E. dilaticollis, could not be differentiated from each other through DNA barcoding. The difference in the DNA sequences between the COI gene of E. integriceps and COI genes of E. maura and E. testudinarius was more than 4%. In the present study DNA barcoding of two Eurygaster species was performed for the first time on E. integriceps, the most dangerous pest in the genus, and E. dilaticollis that only inhabits natural ecosystems. The PCR-RFLP method was developed in this work for the rapid identification of E. integriceps.

  15. Rapid label-free identification of Klebsiella pneumoniae antibiotic resistant strains by the drop-coating deposition surface-enhanced Raman scattering method

    Science.gov (United States)

    Cheong, Youjin; Kim, Young Jin; Kang, Heeyoon; Choi, Samjin; Lee, Hee Joo

    2017-08-01

    Although many methodologies have been developed to identify unknown bacteria, bacterial identification in clinical microbiology remains a complex and time-consuming procedure. To address this problem, we developed a label-free method for rapidly identifying clinically relevant multilocus sequencing typing-verified quinolone-resistant Klebsiella pneumoniae strains. We also applied the method to identify three strains from colony samples, ATCC70063 (control), ST11 and ST15; these are the prevalent quinolone-resistant K. pneumoniae strains in East Asia. The colonies were identified using a drop-coating deposition surface-enhanced Raman scattering (DCD-SERS) procedure coupled with a multivariate statistical method. Our workflow exhibited an enhancement factor of 11.3 × 106 to Raman intensities, high reproducibility (relative standard deviation of 7.4%), and a sensitive limit of detection (100 pM rhodamine 6G), with a correlation coefficient of 0.98. All quinolone-resistant K. pneumoniae strains showed similar spectral Raman shifts (high correlations) regardless of bacterial type, as well as different Raman vibrational modes compared to Escherichia coli strains. Our proposed DCD-SERS procedure coupled with the multivariate statistics-based identification method achieved excellent performance in discriminating similar microbes from one another and also in subtyping of K. pneumoniae strains. Therefore, our label-free DCD-SERS procedure coupled with the computational decision supporting method is a potentially useful method for the rapid identification of clinically relevant K. pneumoniae strains.

  16. Optimum wireless sensor deployment scheme for structural health monitoring: a simulation study

    Science.gov (United States)

    Liu, Chengyin; Fang, Kun; Teng, Jun

    2015-11-01

    With the rapid advancements in smart sensing technology and wireless communication technology, the wireless sensor network (WSN) offers an alternative solution to structural health monitoring (SHM). In WSNs, dense deployment of wireless nodes aids the identification of structural dynamic characteristics, while data transmission is a significant issue since wireless channels typically have a lower bandwidth and a limited power supply. This paper provides a wireless sensor deployment optimization scheme for SHM, in terms of both energy consumption and modal identification accuracy. A spherical energy model is established to formulate the energy consumption within a WSN. The optimal number of sensors and their locations are obtained through solving a multi-objective function with weighting factors on energy consumption and modal identification accuracy using a genetic algorithm (GA). Simulation and comparison results with traditional sensor deployment methods demonstrate the efficiency of the proposed optimization scheme.

  17. Delivery of a national home safety equipment scheme in England: a survey of local scheme leaders.

    Science.gov (United States)

    Mulvaney, C A; Watson, M C; Hamilton, T; Errington, G

    2013-11-01

    Unintentional home injuries sustained by preschool children are a major cause of morbidity in the UK. Home safety equipment schemes may reduce home injury rates. In 2009, the Royal Society for the Prevention of Accidents was appointed as central coordinator of a two-year, £18m national home safety equipment scheme in England. This paper reports the findings from a national survey of all scheme leaders responsible for local scheme delivery. A questionnaire mailed to all local scheme leaders sought details of how the schemes were operated locally; barriers and facilitators to scheme implementation; evaluation of the local scheme and its sustainability. A response rate of 73% was achieved. Health visitors and family support workers played a key role in both the identification of eligible families and performing home safety checks. The majority of local scheme leaders (94.6%) reported that they thought their local scheme had been successful in including those families considered 'harder to engage'. Many scheme leaders (72.4%) reported that they had evaluated the provision of safety equipment in their scheme and over half (56.6%) stated that they would not be able to continue the scheme once funding ceased. Local schemes need support to effectively evaluate their scheme and to seek sustainability funding to ensure the future of the scheme. There remains a lack of evidence of whether the provision of home safety equipment reduces injuries in preschool children.

  18. Rapid and reliable species identification of wild mushrooms by matrix assisted laser desorption/ionization time of flight mass spectrometry (MALDI-TOF MS).

    Science.gov (United States)

    Sugawara, Ryota; Yamada, Sayumi; Tu, Zhihao; Sugawara, Akiko; Suzuki, Kousuke; Hoshiba, Toshihiro; Eisaka, Sadao; Yamaguchi, Akihiro

    2016-08-31

    Mushrooms are a favourite natural food in many countries. However, some wild species cause food poisoning, sometimes lethal, due to misidentification caused by confusing fruiting bodies similar to those of edible species. The morphological inspection of mycelia, spores and fruiting bodies have been traditionally used for the identification of mushrooms. More recently, DNA sequencing analysis has been successfully applied to mushrooms and to many other species. This study focuses on a simpler and more rapid methodology for the identification of wild mushrooms via protein profiling based on matrix-assisted laser desorption/ionization mass spectrometry (MALDI-TOF MS). A preliminary study using 6 commercially available cultivated mushrooms suggested that a more reproducible spectrum was obtained from a portion of the cap than from the stem of a fruiting body by the extraction of proteins with a formic acid-acetonitrile mixture (1 + 1). We used 157 wild mushroom-fruiting bodies collected in the centre of Hokkaido from June to November 2014. Sequencing analysis of a portion of the ribosomal RNA gene provided 134 identifications of mushrooms by genus or species, however 23 samples containing 10 unknown species that had lower concordance rate of the nucleotide sequences in a BLAST search (less than 97%) and 13 samples that had unidentifiable poor or mixed sequencing signals remained unknown. MALDI-TOF MS analysis yielded a reproducible spectrum (frequency of matching score ≥ 2.0 was ≥6 spectra from 12 spectra measurements) for 114 of 157 samples. Profiling scores that matched each other within the database gave correct species identification (with scores of ≥2.0) for 110 samples (96%). An in-house prepared database was constructed from 106 independent species, except for overlapping identifications. We used 48 wild mushrooms that were collected in autumn 2015 to validate the in-house database. As a result, 21 mushrooms were identified at the species level with

  19. Microbial agent detection using near-IR electrophoretic and spectral signatures (MADNESS) for rapid identification in detect-to-warn applications.

    Energy Technology Data Exchange (ETDEWEB)

    Gomez, Anthony Lee; Bambha, Ray P.; VanderNoot, Victoria A.; Fruetel, Julia A.; Renzi, Ronald F.; Krafcik, Karen Lee

    2009-10-01

    Rapid identification of aerosolized biological agents following an alarm by particle triggering systems is needed to enable response actions that save lives and protect assets. Rapid identifiers must achieve species level specificity, as this is required to distinguish disease-causing organisms (e.g., Bacillus anthracis) from benign neighbors (e.g., Bacillus subtilis). We have developed a rapid (1-5 minute), novel identification methodology that sorts intact organisms from each other and particulates using capillary electrophoresis (CE), and detects using near-infrared (NIR) absorbance and scattering. We have successfully demonstrated CE resolution of Bacillus spores and vegetative bacteria at the species level. To achieve sufficient sensitivity for detection needs ({approx}10{sup 4} cfu/mL for bacteria), we have developed fiber-coupled cavity-enhanced absorbance techniques. Using this method, we have demonstrated {approx}two orders of magnitude greater sensitivity than published results for absorbing dyes, and single particle (spore) detection through primarily scattering effects. Results of the integrated CE-NIR system for spore detection are presented.

  20. AADNMR: A Simple Method for Rapid Identification of Bacterial/Mycobacterial Infections in Antibiotic Treated Peritoneal Dialysis Effluent Samples for Diagnosis of Infectious Peritonitis

    CERN Document Server

    Guleria, Anupam; Rawat, Atul; Khetrapal, C L; Prasad, Narayan; Kumar, Dinesh

    2014-01-01

    An efficient method is reported for rapid identification of bacterial or mycobacterial infection in a suspected clinical/biological sample. The method is based on the fact that the ring methylene protons of cyclic fatty acids (constituting the cell membrane of several species of bacteria and mycobacteria) resonate specifically between -0.40 and 0.68 ppm region of the 1H NMR spectrum. These cyclic fatty acids are rarely found in the eukaryotic cell membranes. Therefore, the signals from cyclic ring moiety of these fatty acids can be used as markers (a) for the identification of bacterial and mycobacterial infections and (b) for differential diagnosis of bacterial and fungal infections. However, these microbial fatty acids when present inside the membrane are not easily detectable by NMR owing to their fast T2 relaxation. Nonetheless, the problem can easily be circumvented if these fatty acids become suspended in solution. This has been achieved by abolishing the membrane integrity using broad spectrum antibiot...

  1. CONTRAILS: A tool for rapid identification of transgene integration sites in complex, repetitive genomes using low-coverage paired-end sequencing

    Science.gov (United States)

    Lambirth, Kevin C.; Whaley, Adam M.; Schlueter, Jessica A.; Bost, Kenneth L.; Piller, Kenneth J.

    2015-01-01

    Transgenic crops have become a staple in modern agriculture, and are typically characterized using a variety of molecular techniques involving proteomics and metabolomics. Characterization of the transgene insertion site is of great interest, as disruptions, deletions, and genomic location can affect product selection and fitness, and identification of these regions and their integrity is required for regulatory agencies. Here, we present CONTRAILS (Characterization of Transgene Insertion Locations with Sequencing), a straightforward, rapid and reproducible method for the identification of transgene insertion sites in highly complex and repetitive genomes using low coverage paired-end Illumina sequencing and traditional PCR. This pipeline requires little to no troubleshooting and is not restricted to any genome type, allowing use for many molecular applications. Using whole genome sequencing of in-house transgenic Glycine max, a legume with a highly repetitive and complex genome, we used CONTRAILS to successfully identify the location of a single T-DNA insertion to single base resolution. PMID:26697366

  2. MALDI-TOF MS enables the rapid identification of the major molecular types within the Cryptococcus neoformans/C. gattii species complex.

    Directory of Open Access Journals (Sweden)

    Carolina Firacative

    Full Text Available BACKGROUND: The Cryptococcus neoformans/C. gattii species complex comprises two sibling species that are divided into eight major molecular types, C. neoformans VNI to VNIV and C. gattii VGI to VGIV. These genotypes differ in host range, epidemiology, virulence, antifungal susceptibility and geographic distribution. The currently used phenotypic and molecular identification methods for the species/molecular types are time consuming and expensive. As Matrix-Assisted Laser Desorption Ionization-Time-of-Flight Mass Spectrometry (MALDI-TOF MS offers an effective alternative for the rapid identification of microorganisms, the objective of this study was to examine its potential for the identification of C. neoformans and C. gattii strains at the intra- and inter-species level. METHODOLOGY: Protein extracts obtained via the formic acid extraction method of 164 C. neoformans/C. gattii isolates, including four inter-species hybrids, were studied. RESULTS: The obtained mass spectra correctly identified 100% of all studied isolates, grouped each isolate according to the currently recognized species, C. neoformans and C. gattii, and detected potential hybrids. In addition, all isolates were clearly separated according to their major molecular type, generating greater spectral differences among the C. neoformans molecular types than the C. gattii molecular types, most likely reflecting a closer phylogenetic relationship between the latter. The number of colonies used and the incubation length did not affect the results. No spectra were obtained from intact yeast cells. An extended validated spectral library containing spectra of all eight major molecular types was established. CONCLUSIONS: MALDI-TOF MS is a rapid identification tool for the correct recognition of the two currently recognized human pathogenic Cryptococcus species and offers a simple method for the separation of the eight major molecular types and the detection of hybrid strains within this

  3. Identification by mass spectrometry and automated susceptibility testing from positive bottles: a simple, rapid, and standardized approach to reduce the turnaround time in the management of blood cultures.

    Science.gov (United States)

    Mauri, Carola; Principe, Luigi; Bracco, Silvia; Meroni, Elisa; Corbo, Nicoletta; Pini, Beatrice; Luzzaro, Francesco

    2017-12-06

    Speeding up identification and antimicrobial susceptibility testing (AST) is of foremost importance in the management of blood cultures. Here, we describe a simple, rapid, and standardized approach based on a very short-term incubation on solid medium from positive blood cultures followed by MALDI-TOF mass spectrometry identification and automated AST. The aim of the study was to evaluate the impact in the laboratory practice of this new procedure with respect to that previously used (standard method) by comparing TAT and cumulative percentage of final reports to clinicians. Compared with the standard method, the new procedure gave correct organism identification at genus or species level in 98.4% of monomicrobial samples. AST resulted in 97.7% essential agreement and 98.1% categorical agreement, with 0.9% minor errors, 1.0% major error, and 1.5% very major errors. The mean turnaround time to identification and AST was 61.4 h by using the new method compared to 83.1 h by using standard procedure. Concerning cumulative percentages of final reports, approximately a third of results were available at 48 h from the check-in of the sample when using the new procedure, whereas no final reports were ready at the same time with the standard method. The new procedure allows faster and reliable results using a simple and standardized approach. Thus, it represents an important tool for a more rapid management of blood cultures when molecular methods are not available in the laboratory.

  4. Rapid identification and antimicrobial susceptibility testing of positive blood cultures using MALDI-TOF MS and a modification of the standardised disc diffusion test: a pilot study.

    Science.gov (United States)

    Fitzgerald, C; Stapleton, P; Phelan, E; Mulhare, P; Carey, B; Hickey, M; Lynch, B; Doyle, M

    2016-04-27

    In an era when clinical microbiology laboratories are under increasing financial pressure, there is a need for inexpensive, yet effective, rapid microbiology tests. The aim of this study was to evaluate a novel modification of standard methodology for the identification and antimicrobial susceptibility testing (AST) of pathogens in positive blood cultures, reducing the turnaround time of laboratory results by 24 h. 277 positive blood cultures had a Gram stain performed and were subcultured and incubated at 37°C in a CO2 atmosphere for 4-6 h. Identification of the visible growth was performed using matrix-assisted laser desorption time of flight mass spectrometry (MALDI-TOF MS). Taking a modified approach to the Clinical and Laboratory Standards Institute-standardised AST methodology, an inoculum density of 0.5 McFarland was prepared from the early growth for disc diffusion testing. The standard AST method was also performed on the 18-24 h culture. 96% (n=73/76) of gram-negative organisms were correctly identified by MALDI-TOF MS. Comparative analysis of the rapid and standard AST results showed an overall interpretive category error rate of 7.7% (6.7% minor errors, 0.6% major errors and 0.4% very major errors). 100% of Staphylococcus aureus (n=41) and enterococcus isolates (n=9) were correctly identified after 4-6 h incubation. The overall AST categorical agreement was also 100% for these isolates. An incubation of 4-6 h directly from positive blood cultures allowed for both a rapid species identification and an antimicrobial susceptibility result approximately 24 h earlier than is possible using standard methodology. Published by the BMJ Publishing Group Limited. For permission to use (where not already granted under a licence) please go to http://www.bmj.com/company/products-services/rights-and-licensing/

  5. Development of a rapid diagnostic method for identification of Staphylococcus aureus and antimicrobial resistance in positive blood culture bottles using a PCR-DNA-chromatography method.

    Science.gov (United States)

    Ohshiro, Takeya; Miyagi, Chihiro; Tamaki, Yoshikazu; Mizuno, Takuya; Ezaki, Takayuki

    2016-06-01

    Blood culturing and the rapid reporting of results are essential for infectious disease clinics to obtain bacterial information that can affect patient prognosis. When gram-positive coccoid cells are observed in blood culture bottles, it is important to determine whether the strain is Staphylococcus aureus and whether the strain has resistance genes, such as mecA and blaZ, for proper antibiotic selection. Previous work led to the development of a PCR method that is useful for rapid identification of bacterial species and antimicrobial susceptibility. However, that method has not yet been adopted in community hospitals due to the high cost and methodological complexity. We report here the development of a quick PCR and DNA-chromatography test, based on single-tag hybridization chromatography, that permits detection of S. aureus and the mecA and blaZ genes; results can be obtained within 1 h for positive blood culture bottles. We evaluated this method using 42 clinical isolates. Detection of S. aureus and the resistance genes by the PCR-DNA-chromatography method was compared with that obtained via the conventional identification method and actual antimicrobial susceptibility testing. Our method had a sensitivity of 97.0% and a specificity of 100% for the identification of the bacterial species. For the detection of the mecA gene of S. aureus, the sensitivity was 100% and the specificity was 95.2%. For the detection of the blaZ gene of S. aureus, the sensitivity was 100% and the specificity was 88.9%. The speed and simplicity of this PCR-DNA-chromatography method suggest that our method will facilitate rapid diagnoses. Copyright © 2016 Japanese Society of Chemotherapy and The Japanese Association for Infectious Diseases. Published by Elsevier Ltd. All rights reserved.

  6. Additive operator-difference schemes splitting schemes

    CERN Document Server

    Vabishchevich, Petr N

    2013-01-01

    Applied mathematical modeling isconcerned with solving unsteady problems. This bookshows how toconstruct additive difference schemes to solve approximately unsteady multi-dimensional problems for PDEs. Two classes of schemes are highlighted: methods of splitting with respect to spatial variables (alternating direction methods) and schemes of splitting into physical processes. Also regionally additive schemes (domain decomposition methods)and unconditionally stable additive schemes of multi-component splitting are considered for evolutionary equations of first and second order as well as for sy

  7. The department of veterans affairs nutritional status classification scheme allows for rapid assessment of nutritional status prior to autologous peripheral blood stem cell transplantation and identifies patients at high risk of transplant-related complications.

    Science.gov (United States)

    Toro, Juan J; Haile, David J; Chao, Ju-Hsien; Schneider, Deanna; Jewell, Pamela S; Lee, Shuko; Freytes, César O

    2009-09-01

    The nutritional assessment of patients prior to autologous peripheral blood stem cell transplantation (APBSCT) is labor intensive. A simple method of nutritional assessment prior to APSCT would be extremely helpful, especially if this method could identify patients at high risk of transplant-related complications. The Department of Veterans Affairs (VA) developed a Nutritional Status Classification Scheme (NSCS) to identify nutritionally compromised inpatients rapidly and reliably. The objective of this study was to determine if the use of the VA-NSCS could be utilized as a tool for the evaluation of patients prior to APBSCT and to determine if this tool could be used to identify patients at high risk of transplant-related complications. The nutritional status of 128 patients who underwent APBSCT was assessed by a registered dietician, utilizing the VA-NSCS, upon admission to the hospital and prior to conditioning regimen. Patients with moderately compromised nutritional status pretransplantation experienced a higher incidence of infections, longer duration of diarrhea, and longer length of hospital stay when compared to patients with normal or mildly compromised nutritional status. Our study demonstrates that the VA-NSCS, a simple and inexpensive tool to assess nutritional status, was useful in determining the pretransplant nutritional status of patients with lymphogenous malignancies who underwent APBSCT. In addition, this method was able to identify patients at a higher risk of posttransplant complications. Future studies should be undertaken to determine the optimal method for the nutritional assessment of autologous stem cell transplant candidates.

  8. Surrogate Model Application to the Identification of Optimal Groundwater Exploitation Scheme Based on Regression Kriging Method-A Case Study of Western Jilin Province.

    Science.gov (United States)

    An, Yongkai; Lu, Wenxi; Cheng, Weiguo

    2015-07-30

    This paper introduces a surrogate model to identify an optimal exploitation scheme, while the western Jilin province was selected as the study area. A numerical simulation model of groundwater flow was established first, and four exploitation wells were set in the Tongyu county and Qian Gorlos county respectively so as to supply water to Daan county. Second, the Latin Hypercube Sampling (LHS) method was used to collect data in the feasible region for input variables. A surrogate model of the numerical simulation model of groundwater flow was developed using the regression kriging method. An optimization model was established to search an optimal groundwater exploitation scheme using the minimum average drawdown of groundwater table and the minimum cost of groundwater exploitation as multi-objective functions. Finally, the surrogate model was invoked by the optimization model in the process of solving the optimization problem. Results show that the relative error and root mean square error of the groundwater table drawdown between the simulation model and the surrogate model for 10 validation samples are both lower than 5%, which is a high approximation accuracy. The contrast between the surrogate-based simulation optimization model and the conventional simulation optimization model for solving the same optimization problem, shows the former only needs 5.5 hours, and the latter needs 25 days. The above results indicate that the surrogate model developed in this study could not only considerably reduce the computational burden of the simulation optimization process, but also maintain high computational accuracy. This can thus provide an effective method for identifying an optimal groundwater exploitation scheme quickly and accurately.

  9. Rapid Assay for In Situ Identification of Coagulase-Positive Staphylococci Recovered by Membrane Filtration from Swimming Pool Water

    OpenAIRE

    Klapes, N. Arlene; Vesley, Donald

    1986-01-01

    A rapid, in situ thermonuclease test that identifies colonies of Staphylococcus aureus among staphylococci isolated from swimming pool water by membrane filtration recovery on various selective and differential media is described.

  10. Evaluation of the BD Max StaphSR Assay for Rapid Identification of Staphylococcus aureus and Methicillin-Resistant S. aureus in Positive Blood Culture Broths.

    Science.gov (United States)

    Dalpke, Alexander H; Hofko, Marjeta; Hamilton, Fiona; Mackenzie, Laura; Zimmermann, Stefan; Templeton, Kate

    2015-11-01

    We evaluated the performance of the BD Max StaphSR assay for the direct detection of Staphylococcus aureus from blood culture medium. In a two-center trial, 155 blood cultures from the BD Bactec FX system and 212 from the bioMérieux BacT/Alert system were tested; 170 bottles yielded S. aureus, and all were identified correctly by the BD Max StaphSR assay. The assay required approximately 2.5 h, thus allowing rapid identification of blood cultures flagged positive. Copyright © 2015, American Society for Microbiology. All Rights Reserved.

  11. Rapid identification of microorganisms from positive blood cultures by MALDI-TOF mass spectrometry subsequent to very short-term incubation on solid medium.

    Science.gov (United States)

    Idelevich, E A; Schüle, I; Grünastel, B; Wüllenweber, J; Peters, G; Becker, K

    2014-10-01

    Rapid identification of the causative microorganism is important for appropriate antimicrobial therapy of bloodstream infections. Bacteria from positive blood culture (BC) bottles are not readily available for identification by matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS). Lysis and centrifugation procedures suggested for direct MALDI-TOF MS from positive BCs without previous culture are associated with additional hands-on processing time and costs. Here, we describe an alternative approach applying MALDI-TOF MS from bacterial cultures incubated very briefly on solid medium. After plating of positive BC broth on Columbia blood agar (n = 165), MALDI-TOF MS was performed after 1.5, 2, 3, 4, 5, 6, 7, 8, 12 and (for control) 24 h of incubation until reliable identification to the species level was achieved (score ≥2.0). Mean incubation time needed to achieve species-level identification was 5.9 and 2.0 h for Gram-positive aerobic cocci (GPC, n = 86) and Gram-negative aerobic rods (GNR, n = 42), respectively. Short agar cultures with incubation times ≤2, ≤4, ≤6, ≤8 and ≤12 h yielded species identification in 1.2%, 18.6%, 64.0%, 96.5%, 98.8% of GPC, and in 76.2%, 95.2%, 97.6%, 97.6%, 97.6% of GNR, respectively. Control species identification at 24 h was achieved in 100% of GPC and 97.6% of GNR. Ethanol/formic acid protein extraction performed for an additional 34 GPC isolates cultivated from positive BCs showed further reduction in time to species identification (3.1 h). MALDI-TOF MS using biomass subsequent to very short-term incubation on solid medium allows very early and reliable bacterial identification from positive BCs without additional time and cost expenditure. © 2014 The Authors Clinical Microbiology and Infection © 2014 European Society of Clinical Microbiology and Infectious Diseases.

  12. Restriction fragment length polymorphism of the 5S-rRNA-NTS region: a rapid and precise method for plant identification.

    Science.gov (United States)

    Bertea, Cinzia Margherita; Gnavi, Giorgio

    2012-01-01

    Molecular genetic methods have several advantages over classical morphological and chemical analyses. The genetic method requires genotype instead than phenotype, therefore PCR-based techniques have been widely used for a rapid identification of plant species, varieties and chemotypes. Recently, the molecular discrimination of some higher plant species has been evaluated using sequences of a 5S-rRNA gene spacer region. The variation in the nontranscribed sequence (NTS) region has been used in a number of plant species for studying intraspecific variation, genome evolution, and phylogenetic reconstruction. Here, we describe a rapid method based on the use of the 5S-rRNA-NTS region as a tool for plant DNA fingerprinting, which combines PCR, sequencing and restriction fragment length polymorphism analyses.

  13. Rapid identification of antioxidant compounds of Genista saharae Coss. & Dur. by combination of DPPH scavenging assay and HPTLC-MS.

    Science.gov (United States)

    Meriane, Djamila; Genta-Jouve, Grégory; Kaabeche, Mohamed; Michel, Sylvie; Boutefnouchet, Sabrina

    2014-04-09

    Genista species are sources of antioxidant phenolic compounds such as O- and C-glycosylflavonoids and isoflavonoids. A combination of a DPPH scavenging assay with HPTLC-MS, a fast and efficient method for identification of bioactive compounds, has been applied for evaluation of the radical scavenging activity of metabolites from Genista saharae Coss. & Dur. Different organs collected at various periods have been compared. Identification of antioxidant compounds was obtained by elution of the major DPPH-inhibition zones. The resulting HPTLC-MS analysis under moderately polar conditions, coupled to the DPPH results led to the putative identification of two antioxidant isoflavone aglycones: 3',4',5,7-tetrahydroxyisoflavone (1) and ficuisoflavone (3), whereas polar migration conditions led to the identification of the glycosides 5-methoxy-4',7-trihydroxy-8-glucopyranosylisoflavone (4) and 4',5-dihydroxy-7-methoxyisoflavone-4'-O-β-D-gluco-pyranoside (5). Evaluation of percentage of inhibition of DPPH radical by the purified isoflavone 4 from the root extract showed that it affords a moderate contribution to the total radical scavenging activity of the extract.

  14. Rapid Identification of Staphylococcus aureus Directly from Bactec Blood Culture Broth by the BinaxNOW S. aureus Test

    OpenAIRE

    Qian, Qinfang; Eichelberger, Karen; Kirby, James E.

    2014-01-01

    The BinaxNOW Staphylococcus aureus testing showed sensitivity, specificity, and positive and negative predicative values of 97.6%, 100%, 100%, and 98.4%, respectively, for identification of S. aureus from Bactec blood culture broth. Importantly, the test performed equally well on aerobic and anaerobic culture broth.

  15. Hyphenated chromatographic techniques for the rapid screening and identification of antioxidants in methanolic extracts of pharmaceutically used plants .

    NARCIS (Netherlands)

    Exarchou, V.; Fiamegos, Y.C.; Beek, van T.A.; Nanos, C.G.; Vervoort, J.J.M.

    2006-01-01

    Phytochemical analysis is an important scientific research area, which normally relies on a number of rather laborious and time-consuming techniques for compound identification. Isolation of the ingredients of plant extracts in adequate quantities for spectral and biological analysis was the basis

  16. Rapid Identification of Antioxidant Compounds of Genista saharae Coss. & Dur. by Combination of DPPH Scavenging Assay and HPTLC-MS

    Directory of Open Access Journals (Sweden)

    Djamila Meriane

    2014-04-01

    Full Text Available Genista species are sources of antioxidant phenolic compounds such as O- and C-glycosylflavonoids and isoflavonoids. A combination of a DPPH scavenging assay with HPTLC-MS, a fast and efficient method for identification of bioactive compounds, has been applied for evaluation of the radical scavenging activity of metabolites from Genista saharae Coss. & Dur. Different organs collected at various periods have been compared. Identification of antioxidant compounds was obtained by elution of the major DPPH-inhibition zones. The resulting HPTLC-MS analysis under moderately polar conditions, coupled to the DPPH results led to the putative identification of two antioxidant isoflavone aglycones: 3',4',5,7-tetrahydroxyisoflavone (1 and ficuisoflavone (3, whereas polar migration conditions led to the identification of the glycosides 5-methoxy-4',7-trihydroxy-8-glucopyranosylisoflavone (4 and 4',5-dihydroxy-7-methoxyisoflavone-4'-O-β-D-gluco-pyranoside (5. Evaluation of percentage of inhibition of DPPH radical by the purified isoflavone 4 from the root extract showed that it affords a moderate contribution to the total radical scavenging activity of the extract.

  17. Rapid identification of Salmonella using Hektoen enteric agar and 16s ribosomal DNA probe-gold nanoparticle immunochromatography assay in clinical faecal specimens.

    Science.gov (United States)

    Yeung, C-Y; Liu, C-C; Tseng, Y-T; Tsai, K-C; Hsieh, M-A; Chan, W-T; Liu, H-L; Lee, H-C; Hou, S-Y

    2014-04-01

    A rapid identification of Salmonella, one of the most common foodborne pathogens worldwide, in clinical patients can enable better rational managements and prevent further outbreaks. The traditional immunochromatography using antibody-gold nanoparticles (Ab-AuNPs), such as the home pregnancy test, has been used for the Salmonella detection. In this study, we developed a new and rapid method using DNA probe-AuNPs for the detection of 16s ribosomal DNA of Salmonella. To evaluate the proposed method in clinical specimens, we performed a clinical test by identifying 159 stool samples on Hektoen agar containing black or crystalloid colonies using the method and the VITEK 2 system for confirmation. Eighty of the isolates were correctly identified as Salmonella to achieve 100% sensitivity. Seventy-five samples were correctly identified as non-Salmonella spp., but four were incorrectly identified as Salmonella. The specificity was 94·93%. The assay time is about 30 min after the DNA purification. The time-consuming and labour-intense biochemical tests can be replaced. We demonstrated that this assay is a rapid, convenient and cost-effective tool for Salmonella identification of clinical faecal samples, which is worth for further promotion and clinical use. This is the first application of using 16s ribosomal DNA probe-Au-NPs and immunochromatography on clinical samples. This is the first application of using 16s ribosomal DNA probe-gold nanoparticles and immunochromatography method on clinical samples with sensitivity 100% and specificity 94·93%. The assay time is about 30 min after the DNA purification. We find this assay a rapid, convenient, sensitive and inexpensive tool for Salmonella identification of clinical faecal samples, which is worth further promotion and clinical use and can replace the traditional time-consuming and labour-intense biochemical tests. The potential benefit of this approach is to develop a rapid point-of-care test that provides results while

  18. Reliable and reproducible method for rapid identification of Nocardia species by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry.

    Science.gov (United States)

    Toyokawa, Masahiro; Kimura, Keigo; Nishi, Isao; Sunada, Atsuko; Ueda, Akiko; Sakata, Tomomi; Asari, Seishi

    2013-01-01

    Recently, matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF MS) has been challenged for the identification of Nocardia species. However, the standard ethanol-formic acid extraction alone is insufficient in allowing the membrane proteins of Nocardia species to be ionized by the matrix. We therefore aimed to establish our new extraction method for the MALDI-TOF MS-based identification of Nocardia species isolates. Our modified extraction procedure is through dissociation in 0.5% Tween-20 followed by bacterial heat-inactivation, mechanical breaking of the cell wall by acid-washed glass beads and protein extraction with formic acid and acetonitrile. As reference methods for species identification, full-length 16S rRNA gene sequencing and some phenotypical tests were used. In a first step, we made our own Nocardia database by analyzing 13 strains (13 different species including N. elegans, N. otitidiscaviarum, N. asiatica, N. abscessus, N. brasiliensis, N. thailandica, N. farcinica, N. nova, N. mikamii, N. cyriacigeorgica, N. asteroids, Nocardiopsis alba, and Micromonospora sp.) and registered to the MALDI BioTyper database. Then we established our database. The analysis of 12 challenge strains using the our database gave a 100% correct identification, including 8 strains identified to the species level and 4 strains to the genus level (N. elegans, N. nova, N. farcinica, Micromonospora sp.) according to the manufacture's log score specifications. In the estimation of reproducibility of our method intended for 4 strains, both within-run and between-run reproducibility were excellent. These data indicates that our method for rapid identification of Nocardia species is with reliability, reproducibility and cost effective.

  19. An enzymatic deglycosylation scheme enabling identification of core fucosylated N-glycans and O-glycosylation site mapping of human plasma proteins

    DEFF Research Database (Denmark)

    Hägglund, Per; Matthiesen, R.; Elortza, F.

    2007-01-01

    and N-acetyl-β-glucosaminidase) are also included. The two strategies were here applied to identify 103 N-glycosylation sites in the Cohn IV fraction of human plasma. In addition, Endo D/H digestion uniquely enabled identification of 23 fucosylated N-glycosylation sites. Several O-glycosylated peptides......, thereby reducing the complexity and facilitating glycosylation site determinations. Here, we have used two different enzymatic deglycosylation strategies for N-glycosylation site analysis. (1) Removal of entire N-glycan chains by peptide- N-glycosidase (PNGase) digestion, with concomitant deamidation...... of the released asparagine residue. The reaction is carried out in H218O to facilitate identification of the formerly glycosylated peptide by incorporatation of 18O into the formed aspartic acid residue. (2) Digestion with two endo-β- N-acetylglucosaminidases (Endo D and Endo H) that cleave the glycosidic bond...

  20. Room temperature DNA preservation of soft tissue for rapid DNA extraction: an addition to the disaster victim identification investigators toolkit?

    Science.gov (United States)

    Graham, E A M; Turk, E E; Rutty, G N

    2008-01-01

    In mass fatality incidents, for example following a vehicle accident or terrorist event, severe fragmentation of bodies may occur, making identification by the use of traditional techniques such as fingerprinting or odontology difficult. In such situations DNA profiling can be employed for individualization and re-association of fragmented remains. As at times disrupted soft tissue may be the predominate tissue type requiring identification and re-association. We have investigated the use of two buffer solutions for preservation of soft tissue samples that may be collected during such investigations, when buccal cells, blood samples or teeth or bone may not be available. Both buffer solutions have shown sufficient DNA preservation over a 12-month period of storage at room temperature to allow for DNA profiling to be successfully performed when 5-1000 mg muscle tissue was stored in each solution.

  1. Applying characteristic fragment filtering for rapid detection and identification of ingredients in rhubarb by HPLC coupled with linear ion trap-Orbitrap mass spectrometry.

    Science.gov (United States)

    Wang, Qing; Lu, Zhiwei; Zhang, Li; Zhang, Qingqing; Wang, Meiling; Zhao, Huizhen; Liu, Yuehong; Fu, Shuang; Huang, Zhenghai; Xie, Ziye; Yu, Honghong; Zhang, Zhixin; Gao, Xiaoyan

    2017-07-01

    Chemical characteristic fragment filtering in MSn chromatograms was proposed to detect and identify the components in rhubarb rapidly using high-performance liquid chromatography coupled with linear ion trap-Orbitrap mass spectrometry. Characteristic fragments consist of diagnostic ions and neutral loss fragments. Characteristic fragment filtering is a postacquisition data mining method for the targeted screening of groups with specific structures, including three steps: first, in order to comprehensively summarize characteristic fragments for global identification of the ingredients in rhubarb, representative authentic standards of dominant chemical categories contained in rhubarb were chosen, from which fragmentation rules and a characteristic fragments schedule were proposed; second, characteristic fragment filtering was used to rapidly recognize analogous skeletons; finally, combined with retention time, accurate mass, characteristic fragments, and previous literature, the structures of the filtered compounds were identified or tentatively characterized. As a result, a total of 271 compounds were detected and identified in rhubarb, including 34 anthraquinones, 83 anthrones, 46 tannins, 17 stilbenes, 24 phenylbutanones, 26 acylglucosides, 26 chromones, and 15 other compounds, 69 of which are potentially new compounds. The proposed characteristic fragment filtering strategy would be a reference for the large-scale detection and identification of the ingredients of herbal medicines. © 2017 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  2. A rapid, low-cost, and microfluidic chip-based system for parallel identification of multiple pathogens related to clinical pneumonia.

    Science.gov (United States)

    Huang, Guoliang; Huang, Qin; Xie, Lan; Xiang, Guangxin; Wang, Lei; Xu, Hui; Ma, Li; Luo, Xianbo; Xin, Juan; Zhou, Xinying; Jin, Xiangyu; Zhang, Lei

    2017-07-25

    An air-insulated microfluidic chip was designed for the automatic centrifugal distribution of samples to 24-test cells, enabling the parallel identification of multiple clinical pneumonia-related pathogens in 1.45-μL reactions without cross-contamination in 45 min. A portable nucleic acid analyzer that integrates mechanical, confocal optical, electronic, and software functions was also developed to collect fluorescence data in a Ø3 mm imaging field near the optical diffraction limit for highly sensitive fluorescence detection of nucleic acid amplification in real time. This microfluidic chip-based portable nucleic acid analyzer could detect low abundance nucleic acids present at as few as 10 copies. In a blinded experiment, specific identification of Mycoplasma pneumoniae, Staphylococcus aureus, and methicillin-resistant S. aureus was achieved with 229 clinical patient sputum samples. The total coincidence rate of our system and traditional RT-PCR with an ABI 7500 was 99.56%. Four samples accounting for the 0.44% inconformity were retested by gene sequencing, revealing that our system reported the correct results. This novel microfluidic chip-based detection system is cost-effective, rapid, sensitive, specific, and has a relatively high throughput for parallel identification, which is especially suitable for resource-limited facilities/areas and point-of-care testing.

  3. Internal transcribed spacer sequence-based rapid molecular identification of Prototheca zopfii and Prototheca blaschkeae directly from milk of infected cows.

    Science.gov (United States)

    Marques, S; Huss, V A R; Pfisterer, K; Grosse, C; Thompson, G

    2015-05-01

    The increasing incidence of rare mastitis-causing pathogens has urged the implementation of fast and efficient diagnostic and control measures. Prototheca algae are known to be associated with diseases in humans and animals. In the latter, the most prevalent form of protothecosis is bovine mastitis with Prototheca zopfii and Prototheca blaschkeae representing the most common pathogenic species. These nonphotosynthetic and colorless green algae are ubiquitous in different environments and are widely resistant against harmful conditions and antimicrobials. Hence, the association of Prototheca with bovine mastitis represents a herd problem, requiring fast and easy identification of the infectious agent. The purpose of this study was to develop a reliable and rapid method, based on the internal transcribed spacer (ITS) sequences of ribosomal DNA, for molecular identification and discrimination between P. zopfii and P. blaschkeae in bovine mastitic milk. The complete ITS sequences of 32 Prototheca isolates showed substantial interspecies but moderate intraspecies variability facilitating the design of species-specific PCR amplification primers. The species-specific PCR was successfully applied to the identification of P. zopfii and P. blaschkeae directly from milk samples. The intraspecific ITS phylogeny was compared for each species with the geographical distribution of the respective Prototheca isolates, but no significant correlation was found. Copyright © 2015 American Dairy Science Association. Published by Elsevier Inc. All rights reserved.

  4. Rapid identification of carbapenemase-producing Enterobacteriaceae, Pseudomonas aeruginosa and Acinetobacter baumannii using a modified Carba NP test

    Directory of Open Access Journals (Sweden)

    S. Bakour

    2015-09-01

    Full Text Available Biochemical tests have been previously developed to identify carbapenemase-producing Enterobacteriaceae, Pseudomonas spp. (Carba NP test and Acinetobacter spp. (CarbAcineto NP test. We evaluated a modified Carba NP test to detect carbapenemase production in Enterobacteriaceae, Pseudomonas and Acinetobacter species using a single protocol with rapid results and found good reliability and speed.

  5. A sensitive and a rapid multiplex polymerase chain reaction for the identification of Candida species in concentrated oral rinse specimens in patients with diabetes.

    Science.gov (United States)

    Sampath, Asanga; Weerasekera, Manjula; Gunasekara, Chinthika; Dilhari, Ayomi; Bulugahapitiya, Uditha; Fernando, Neluka

    2017-03-01

    Oral candidiasis is being frequently recognized in patients with diabetes, and is associated with multiple pathogens including Candida albicans, Candida parapsilosis, Candida glabrata and Candida tropicalis. The aim of this study was to evaluate a usefulness of a Multiplex Polymerase Chain Reaction as a rapid diagnostic tool for identification of four oral Candida pathogens in patients with diabetes. A multiplex PCR was optimized to identify four Candida species in concentrated oral rinse samples. Common reverse primer, ITS4 and four species-specific forward primers targeting ITS1 and ITS2 regions of yeast genome were used. Species-specific single amplicon were detected by agarose gel electrophoresis. Performance efficacy of multiplex PCR was compared with phenotypic identification. Out of 100 oral rinse samples, 72 were culture positive and of these 43 were at risk of oral Candida infection (>600cfu/ml). Multiple Candida species including C. albicans, C. parapsilosis and C. tropicalis were identified in 22 samples which had risk of oral Candida infection. In total, 85 patients were positive for Candida by multiplex PCR and of them 49 had multiple Candida species. All 43 colonized specimens were also positive by multiplex PCR. C. albicans was the most predominant organism (75/85) followed by C. parapsilosis (47/85), C. tropicalis (17/85) and C. glabrata (6/85). In specimens with multiple species, the two most common organisms were C. albicans and C. parapsilosis. Multiplex PCR yielded a sensitivity of 10 Candida cells/ml of oral rinse sample. Multiplex PCR is found to be rapid, sensitive and specific than phenotypic identification methods in discriminating multiple Candida species in oral rinse specimens.

  6. Scheme Program Documentation Tools

    DEFF Research Database (Denmark)

    Nørmark, Kurt

    2004-01-01

    This paper describes and discusses two different Scheme documentation tools. The first is SchemeDoc, which is intended for documentation of the interfaces of Scheme libraries (APIs). The second is the Scheme Elucidator, which is for internal documentation of Scheme programs. Although the tools...

  7. A Multipurpose Key Agreement Scheme in Ubiquitous Computing Environments

    Directory of Open Access Journals (Sweden)

    Chin-Chen Chang

    2015-01-01

    Full Text Available Due to the rapid advancement of cryptographic techniques, the smart card has recently become a popular device because it is capable of storing and computing essential information with such properties as tamper resistance. However, many service providers must satisfy the user’s desire to be able to access services anytime and anywhere with the smart card computing devices. Therefore, multipurpose smart cards have become very popular identification tokens. In 2011, Wang et al. proposed an authentication and key agreement scheme for smart card use. Even so, two drawbacks still exist; that is, (1 the security requirement of mutual authentication has not been satisfied and (2 the authentication scheme cannot be used for multipurpose smart cards. In this paper, we propose an efficient and secure multipurpose, authenticated, key agreement scheme in which the user is required to register only once and can be authenticated without any registration center. Furthermore, the proposed scheme can be used in ubiquitous environments because of its low computation and communication overhead.

  8. Bacterial rapid identification with matrix assisted laser desorption/ionization time-of-flight mass spectrometry: development of an 'in-house method' and comparison with Bruker Sepsityper(®) kit.

    Science.gov (United States)

    Frédéric Ric, S; Antoine, M; Bodson, A; Lissoir, B

    2015-10-01

    The objective of this study was to compare an in-house matrix-assisted laser desorption ionization with time of flight (MALDI-TOF) method and a commercial MALDI-TOF kit (Sepsityper(®) kit) for direct bacterial identification in positive blood cultures. We also evaluated the time saved and the cost associated with the rapid identification techniques. We used the BACTEC(®) automated system for detecting positive blood cultures. Direct identification using Sepsityper kit and the in-house method were compared with conventional identification by MALDI-TOF using pure bacterial culture on the solid phase. We also evaluated different cut-off scores for rapid bacterial identification. In total, 127 positive blood vials were selected. The rate of rapid identification with the MALDI Sepsityper kit was 25.2% with the standard cut-off and 33.9% with the enlarged cut-off, while the results for the in-house method were 44.1 and 61.4%, respectively. Error rates with the enlarged cut-off were 6.98 (n = 3) and 2.56% (n = 2) for Sepsityper and the in-house method, respectively. Identification rates were higher for gram-negative bacteria. Direct bacterial identification succeeded in supplying rapid identification of the causative organism in cases of sepsis. The time taken to obtain a result was nearly 24  hours shorter for the direct bacterial identification methods than for conventional MALDI-TOF on solid phase culture. Compared with the Sepsityper kit, the in-house method offered better results and fewer errors, was more cost-effective and easier to use.

  9. Prospective assessment of FilmArray® technology for the rapid identification of yeast isolated from blood cultures.

    Science.gov (United States)

    Desoubeaux, Guillaume; Franck-Martel, Claire; Bailly, Éric; Le Brun, Cécile; Gyan, Emmanuel; Goudeau, Alain; Chandenier, Jacques; Lanotte, Philippe

    2014-11-01

    We prospectively assessed the ability of FilmArray® device to identify fungal species involved in bloodstream infections. It succeeded in identifying 85.7% of isolates. The automated readout of results enabled the rapid initiation of appropriate antifungal therapy. Thus, FilmArray® appeared as a reliable alternative diagnostic method for the most common yeast-like species. Copyright © 2014 Elsevier B.V. All rights reserved.

  10. Rapid identification of thousands of copperhead snake (Agkistrodon contortrix) microsatellite loci from modest amounts of 454 shotgun genome sequence

    OpenAIRE

    Todd A. Castoe*; Poole, Alexander W.; Gu, Wanjun; A.P. Jason de Koning*; Daza, Juan M.; Eric N. Smith; Pollock, David D.

    2009-01-01

    Optimal integration of next-generation sequencing into mainstream research requires reevaluation of how problems can be reasonably overcome and what questions can be asked. One potential application is the rapid acquisition of genomic information to identify microsatellite loci for evolutionary, population genetic and chromosome linkage mapping research on non-model and not previously sequenced organisms. Here, we report on results using high-throughput sequencing to obtain a large number of ...

  11. Rapid identification of a Mycobacterium tuberculosis full genetic drug resistance profile through whole genome sequencing directly from sputum

    Directory of Open Access Journals (Sweden)

    Camus Nimmo

    2017-09-01

    Discussion: Compared to rapid molecular tests (which can only examine a limited number of mutations and WGS of culture isolates (which requires a culture step, WGS directly from sputum can quickly generate a complete genetic drug resistance profile. In this case, WGS altered the clinical management of drug-resistant tuberculosis and demonstrated potential for guiding individualized drug treatment where second-line drug resistance is common.

  12. Rapid antimicrobial susceptibility test for identification of new therapeutics and drug combinations against multidrug-resistant bacteria

    OpenAIRE

    Sun, Wei; Weingarten, Rebecca A; Xu, Miao; Southall, Noel; Dai, Sheng; Shinn, Paul; Sanderson, Philip E; Williamson, Peter R; Frank, Karen M; Zheng, Wei

    2016-01-01

    Current antimicrobial susceptibility testing has limited screening capability for identifying empirical antibiotic combinations to treat severe bacterial infections with multidrug-resistant (MDR) organisms. We developed a new antimicrobial susceptibility assay using automated ultra-high-throughput screen technology in combination with a simple bacterial growth assay. A rapid screening of 5170 approved drugs and other compounds identified 25 compounds with activities against MDR Klebsiella pne...

  13. Direct identification of bacteria in positive blood cultures: comparison of two rapid methods, FilmArray and mass spectrometry.

    Science.gov (United States)

    Rand, Kenneth H; Delano, John P

    2014-07-01

    We evaluated the accuracy and performance of the FilmArray Direct from Positive Blood Culture system (BCID) (BioFire Diagnostics, Salt Lake City, UT, USA) and the VITEK Mass Spectrometry System (Vitek MS; bioMerieux, Durham, NC, USA) to identify bacterial isolates from 161 positive blood culture bottles. The BCID uses multiplex PCR to identify 90-95% of common isolates to the genus or species/complex level as well as mecA, Van A/B, and bla(KPC) genes in approximately 1 hour. Of 151 monomicrobic isolates, the FilmArray correctly identified 48/49 (98%) to the genus and 84/84 (100%) to the species/complex level, while 18/151 (12%) gave no identification, as expected from the database. Mass spectrometry correctly identified 142/151 (94%) monomicrobic cultures to the genus level, 137/151 (91%) to the species level, with only 8/151(5%) giving no identification. Although mass spectrometry has a much larger database, the filtration system was cumbersome in contrast to the 3-5 minutes hands-on-time for the BCID. Copyright © 2014 Elsevier Inc. All rights reserved.

  14. A rapid interleukin-6 bedside test for the identification of intra-amniotic inflammation in preterm labor with intact membranes

    Science.gov (United States)

    Chaemsaithong, Piya; Romero, Roberto; Korzeniewski, Steven J.; Martinez-Varea, Alicia; Dong, Zhong; Yoon, Bo Hyun; Hassan, Sonia S.; Chaiworapongsa, Tinnakorn; Yeo, Lami

    2016-01-01

    Abstract Objective: Preterm birth is associated with 5–18% of pregnancies and is the leading cause of neonatal morbidity and mortality. Amniotic fluid (AF) interleukin-6 (IL-6) is a key cytokine for the identification of intra-amniotic inflammation, and patients with an elevated AF IL-6 are at risk for impending preterm delivery. However, results of the conventional method of measurement (enzyme-linked immunosorbent assay; ELISA) are usually not available in time to inform care. The objective of this study was to determine whether a point of care (POC) test or lateral-flow-based immunoassay for measurement of AF IL-6 concentrations can identify patients with intra-amniotic inflammation and/or infection and those destined to deliver spontaneously before term among women with preterm labor and intact membranes. Methods: One-hundred thirty-six women with singleton pregnancies who presented with symptoms of preterm labor and underwent amniocentesis were included in this study. Amniocentesis was performed at the time of diagnosis of preterm labor. AF Gram stain and AF white blood cell counts were determined. Microbial invasion of the amniotic cavity (MIAC) was defined according to the results of AF culture (aerobic and anaerobic as well as genital mycoplasmas). AF IL-6 concentrations were determined by both lateral flow-based immunoassay and ELISA. The primary outcome was intra-amniotic inflammation, defined as AF ELISA IL-6 ≥ 2600 pg/ml. Results: (1) AF IL-6 concentrations determined by a POC test have high sensitivity (93%), specificity (91%) and a positive likelihood ratio of 10 for the identification of intra-amniotic inflammation by using a threshold of 745 pg/ml; (2) the POC test and ELISA for IL-6 perform similarly in the identification of MIAC, acute inflammatory lesions of placenta and patients at risk of impending spontaneous preterm delivery. Conclusion: A POC AF IL-6 test can identify intra-amniotic inflammation in women who present with preterm

  15. Rapid species identification of "Streptococcus milleri" strains by line blot hybridization: identification of a distinct 16S rRNA population closely related to Streptococcus constellatus.

    Science.gov (United States)

    Jacobs, J A; Schot, C S; Bunschoten, A E; Schouls, L M

    1996-01-01

    A collection of 399 "Streptococcus milleri" strains were identified to the species level by the use of a line blot assay. Their PCR-amplified partial 16S rRNA gene sequences were hybridized with species-specific 5'-biotinylated oligonucleotide probes homologous to the bp 213 to 231 regions of the 16S rRNA gene sequences of the type strains Streptococcus anginosus ATCC 33397, Streptococcus constellatus ATCC 27823, and Streptococcus intermedius ATCC 27335. The hybridization results were compared with the reference phenotypic identification method data (R. A. Whiley, H. Fraser, J. M. Hardie, and D. Beighton, J. Clin. Microbiol. 28:1497-1501, 1990). Most strains (357 of 399 [89.5%]) reacted unambiguously with only one probe. However, 42 of the 399 strains (10.5%) reacted with both the S. constellatus- and S. intermedius-specific probes; 41 of them were phenotypically identified as S. constellatus. These dually reactive strains hybridized with a 5'-biotinylated probe based on the bp 213 to 231 region of the 16S rRNA gene sequence of one of two species. Analysis of the 5' ends of the 16S rRNA gene sequences (487 bp) demonstrated that the dually reactive strains represent a distinct rRNA population sharing 98.1% sequence similarity with S. constellatus. Phenotypic consistency between the dually reactive strains and the S. constellatus strains was not demonstrated. Line blot hybridization proved to be a simple and inexpensive method to screen large numbers of strains for genetic relatedness, and it allowed the detection of a distinct 16S rRNA type within the "S. milleri" group. PMID:8784576

  16. Rapid sample preparation for detection and identification of avian influenza virus from chicken faecal samples using magnetic bead microsystem

    DEFF Research Database (Denmark)

    Dhumpa, Raghuram; Bu, Minqiang; Handberg, Kurt

    2010-01-01

    -PCR is a sensitive method for detection of AIV, it requires sample preparation including separation and purification of AIV and concentrate viral RNA. It is laborious and complex process especially for diagnosis using faecal sample. In this study, magnetic beads were used for immunoseparation of AIV in chicken...... faecal sample by a magnetic microsystem. Using this system, all the 16 hemagglutinin (H) and 9 neuraminidase (N) subtypes of AIV were separated and detected in spiked faecal samples using RT-PCR, without an RNA extraction step. This rapid sample preparation method can be integrated with a total analysis...

  17. Rapid detection and identification of Brugia malayi, B. pahangi, and Dirofilaria immitis by high-resolution melting assay.

    Science.gov (United States)

    Wongkamchai, Sirichit; Monkong, Nuntiya; Mahannol, Pakpimom; Taweethavonsawat, Piyanan; Loymak, Sumat; Foongladda, Suporn

    2013-01-01

    Human lymphatic filariasis is caused by filarial worms such as Brugia malayi for which the major reservoir is domestic cats. However, domestic cats or dogs also carry nonhuman filaria such as Brugia pahangi and Dirofilaria immitis. We have developed a single-tube, real-time PCR with a high-resolution melting (HRM) analysis assay for detection and identification of B. malayi, B. pahangi, and D. immitis in blood samples. The designated primer pair in the PCR can amplify a 114-bp region of mitochondrial 12S rRNA genes of these filarial worms. Subsequently, the HRM assay showed a specific melting temperature for each species. The assay showed the highest sensitivity and specificity in comparison with DNA sequences after assessment with 34 cat and 14 dog blood samples. This assay could be helpful for epidemiological studies of reservoirs and vectors.

  18. A rapid genetic assay for the identification of the most common Pocillopora damicornis genetic lineages on the Great Barrier Reef.

    Directory of Open Access Journals (Sweden)

    Gergely Torda

    Full Text Available Pocillopora damicornis (Linnaeus, 1758; Scleractinia, Pocilloporidae has recently been found to comprise at least five distinct genetic lineages in Eastern Australia, some of which likely represent cryptic species. Due to similar and plastic gross morphology of these lineages, field identification is often difficult. Here we present a quick, cost effective genetic assay as well as three novel microsatellite markers that distinguish the two most common lineages found on the Great Barrier Reef. The assay is based on PCR amplification of two regions within the mitochondrial putative control region, which show consistent and easily identifiable fragment size differences for the two genetic lineages after Alu1 restriction enzyme digestion of the amplicons.

  19. Chemometric Analysis of Lavender Essential Oils Using Targeted and Untargeted GC-MS Acquired Data for the Rapid Identification and Characterization of Oil Quality.

    Science.gov (United States)

    Beale, David J; Morrison, Paul D; Karpe, Avinash V; Dunn, Michael S

    2017-08-11

    Standard raw material test methods such as the ISO Standard 11024 are focused on the identification of lavender oil and not the actual class/quality of the oil. However, the quality of the oil has a significant effect on its price at market. As such, there is a need for raw material tests to identify not only the type of oil but its quality. This paper describes two approaches to rapidly identifying and classifying lavender oil. First, the ISO Standard 11024 test method was evaluated in order to determine its suitability to assess lavender oil quality but due to its targeted and simplistic approach, it has the potential to miss classify oil quality. Second, utilizing the data generated by the ISO Standard 11024 test methodology, an untargeted chemometric predicative model was developed in order to rapidly assess and characterize lavender oils (Lavandulaangustifolia L.) for geographical/environmental adulteration that impact quality. Of the 170 compounds identified as per the ISO Standard 11024 test method utilizing GC-MS analyses, 15 unique compounds that greatly differentiate between the two classes of lavender were identified. Using these 15 compounds, a predicative multivariate chemometric model was developed that enabled lavender oil samples to be reliably differentiated based on quality. A misclassification analysis was performed and it was found that the predictions were sound (100% matching rate). Such an approach will enable producers, distributers, suppliers and manufactures to rapidly screen lavender essential oil. The authors concede that the validation and implementation of such an approach is more difficult than a conventional chromatographic assay. However, the rapid, reliable and less problematic screening is vastly superior and easily justifies any early implementation validation difficulties and costs.

  20. Rapid identification of pearl powder from Hyriopsis cumingii by Tri-step infrared spectroscopy combined with computer vision technology

    Science.gov (United States)

    Liu, Siqi; Wei, Wei; Bai, Zhiyi; Wang, Xichang; Li, Xiaohong; Wang, Chuanxian; Liu, Xia; Liu, Yuan; Xu, Changhua

    2018-01-01

    Pearl powder, an important raw material in cosmetics and Chinese patent medicines, is commonly uneven in quality and frequently adulterated with low-cost shell powder in the market. The aim of this study is to establish an adequate approach based on Tri-step infrared spectroscopy with enhancing resolution combined with chemometrics for qualitative identification of pearl powder originated from three different quality grades of pearls and quantitative prediction of the proportions of shell powder adulterated in pearl powder. Additionally, computer vision technology (E-eyes) can investigate the color difference among different pearl powders and make it traceable to the pearl quality trait-visual color categories. Though the different grades of pearl powder or adulterated pearl powder have almost identical IR spectra, SD-IR peak intensity at about 861 cm- 1 (v2 band) exhibited regular enhancement with the increasing quality grade of pearls, while the 1082 cm- 1 (v1 band), 712 cm- 1 and 699 cm- 1 (v4 band) were just the reverse. Contrastly, only the peak intensity at 862 cm- 1 was enhanced regularly with the increasing concentration of shell powder. Thus, the bands in the ranges of (1550-1350 cm- 1, 730-680 cm- 1) and (830-880 cm- 1, 690-725 cm- 1) could be exclusive ranges to discriminate three distinct pearl powders and identify adulteration, respectively. For massive sample analysis, a qualitative classification model and a quantitative prediction model based on IR spectra was established successfully by principal component analysis (PCA) and partial least squares (PLS), respectively. The developed method demonstrated great potential for pearl powder quality control and authenticity identification in a direct, holistic manner.

  1. Rapid identification of pearl powder from Hyriopsis cumingii by Tri-step infrared spectroscopy combined with computer vision technology.

    Science.gov (United States)

    Liu, Siqi; Wei, Wei; Bai, Zhiyi; Wang, Xichang; Li, Xiaohong; Wang, Chuanxian; Liu, Xia; Liu, Yuan; Xu, Changhua

    2018-01-15

    Pearl powder, an important raw material in cosmetics and Chinese patent medicines, is commonly uneven in quality and frequently adulterated with low-cost shell powder in the market. The aim of this study is to establish an adequate approach based on Tri-step infrared spectroscopy with enhancing resolution combined with chemometrics for qualitative identification of pearl powder originated from three different quality grades of pearls and quantitative prediction of the proportions of shell powder adulterated in pearl powder. Additionally, computer vision technology (E-eyes) can investigate the color difference among different pearl powders and make it traceable to the pearl quality trait-visual color categories. Though the different grades of pearl powder or adulterated pearl powder have almost identical IR spectra, SD-IR peak intensity at about 861cm-1 (v2 band) exhibited regular enhancement with the increasing quality grade of pearls, while the 1082cm-1 (v1 band), 712cm-1 and 699cm-1 (v4 band) were just the reverse. Contrastly, only the peak intensity at 862cm-1 was enhanced regularly with the increasing concentration of shell powder. Thus, the bands in the ranges of (1550-1350cm-1, 730-680cm-1) and (830-880cm-1, 690-725cm-1) could be exclusive ranges to discriminate three distinct pearl powders and identify adulteration, respectively. For massive sample analysis, a qualitative classification model and a quantitative prediction model based on IR spectra was established successfully by principal component analysis (PCA) and partial least squares (PLS), respectively. The developed method demonstrated great potential for pearl powder quality control and authenticity identification in a direct, holistic manner. Copyright © 2017. Published by Elsevier B.V.

  2. Effect of Electrode Geometry on the Classification Performance of Rapid Evaporative Ionization Mass Spectrometric (REIMS) Bacterial Identification.

    Science.gov (United States)

    Bodai, Zsolt; Cameron, Simon; Bolt, Frances; Simon, Daniel; Schaffer, Richard; Karancsi, Tamas; Balog, Julia; Rickards, Tony; Burke, Adam; Hardiman, Kate; Abda, Julia; Rebec, Monica; Takats, Zoltan

    2018-01-01

    The recently developed automated, high-throughput monopolar REIMS platform is suited for the identification of clinically important microorganisms. Although already comparable to the previously reported bipolar forceps method, optimization of the geometry of monopolar electrodes, at the heart of the system, holds the most scope for further improvements to be made. For this, sharp tip and round shaped electrodes were optimized to maximize species-level classification accuracy. Following optimization of the distance between the sample contact point and tube inlet with the sharp tip electrodes, the overall cross-validation accuracy improved from 77% to 93% in negative and from 33% to 63% in positive ion detection modes, compared with the original 4 mm distance electrode. As an alternative geometry, round tube shaped electrodes were developed. Geometry optimization of these included hole size, number, and position, which were also required to prevent plate pick-up due to vacuum formation. Additional features, namely a metal "X"-shaped insert and a pin in the middle were included to increase the contact surface with a microbial biomass to maximize aerosol production. Following optimization, cross-validation scores showed improvement in classification accuracy from 77% to 93% in negative and from 33% to 91% in positive ion detection modes. Supervised models were also built, and after the leave 20% out cross-validation, the overall classification accuracy was 98.5% in negative and 99% in positive ion detection modes. This suggests that the new generation of monopolar REIMS electrodes could provide substantially improved species level identification accuracies in both polarity detection modes. Graphical abstract.

  3. Flow Injection Analysis with Electrochemical Detection for Rapid Identification of Platinum-Based Cytostatics and Platinum Chlorides in Water

    Directory of Open Access Journals (Sweden)

    Marketa Kominkova

    2014-02-01

    Full Text Available Platinum-based cytostatics, such as cisplatin, carboplatin or oxaliplatin are widely used agents in the treatment of various types of tumors. Large amounts of these drugs are excreted through the urine of patients into wastewaters in unmetabolised forms. This phenomenon leads to increased amounts of platinum ions in the water environment. The impacts of these pollutants on the water ecosystem are not sufficiently investigated as well as their content in water sources. In order to facilitate the detection of various types of platinum, we have developed a new, rapid, screening flow injection analysis method with electrochemical detection (FIA-ED. Our method, based on monitoring of the changes in electrochemical behavior of analytes, maintained by various pH buffers (Britton-Robinson and phosphate buffer and potential changes (1,000, 1,100 and 1,200 mV offers rapid and cheap selective determination of platinum-based cytostatics and platinum chlorides, which can also be present as contaminants in water environments.

  4. Rapid identification of thousands of copperhead snake (Agkistrodon contortrix) microsatellite loci from modest amounts of 454 shotgun genome sequence.

    Science.gov (United States)

    Castoe, Todd A; Poole, Alexander W; Gu, Wanjun; Jason de Koning, A P; Daza, Juan M; Smith, Eric N; Pollock, David D

    2010-03-01

    Optimal integration of next-generation sequencing into mainstream research requires re-evaluation of how problems can be reasonably overcome and what questions can be asked. One potential application is the rapid acquisition of genomic information to identify microsatellite loci for evolutionary, population genetic and chromosome linkage mapping research on non-model and not previously sequenced organisms. Here, we report on results using high-throughput sequencing to obtain a large number of microsatellite loci from the venomous snake Agkistrodon contortrix, the copperhead. We used the 454 Genome Sequencer FLX next-generation sequencing platform to sample randomly ∼27 Mbp (128 773 reads) of the copperhead genome, thus sampling about 2% of the genome of this species. We identified microsatellite loci in 11.3% of all reads obtained, with 14 612 microsatellite loci identified in total, 4564 of which had flanking sequences suitable for polymerase chain reaction primer design. The random sequencing-based approach to identify microsatellites was rapid, cost-effective and identified thousands of useful microsatellite loci in a previously unstudied species. © 2009 Blackwell Publishing Ltd.

  5. Laser Induced breakdown spectroscopy: A rapid tool for the identification and quantification of minerals in cucurbit seeds.

    Science.gov (United States)

    Singh, Jyotsana; Kumar, Rohit; Awasthi, Shikha; Singh, Vinti; Rai, A K

    2017-04-15

    Laser-induced breakdown spectroscopy (LIBS) was investigated to estimate the viability as a simple and rapid method for analysis of nutrient elements in seed kernels of cucurbits. LIBS spectra were recorded in the range of 200-975nm by using Q-switched Nd:YAG laser at 532nm (4ns, 10Hz) attached to echelle spectrometer with intensified charged coupled device (ICCD). The spectral analysis revealed the presence of several elements like C, O, N, Mg, Ca, Na and K in seeds. The quantification of elements (Mg, Ca, Na and K) through LIBS was done using calibration curve method in which all calibration curve shows good linearity (r>0.95). The result obtained through LIBS was in reasonable agreement with that obtained through atomic absorption spectroscopy (AAS). Principal Component Analysis (PCA) was also applied to the LIBS data for rapid categorization of seed samples belonging to same species although samples have similar nutrient elements. Copyright © 2016 Elsevier Ltd. All rights reserved.

  6. The use of immunochromatographic rapid test for soft tissue remains identification in order to distinguish between human and non-human origin.

    Science.gov (United States)

    Gascho, Dominic; Morf, Nadja V; Thali, Michael J; Schaerli, Sarah

    2017-05-01

    Clear identification of soft tissue remains as being of non-human origin may be visually difficult in some cases e.g. due to decomposition. Thus, an additional examination is required. The use of an immunochromatographic rapid tests (IRT) device can be an easy solution with the additional advantage to be used directly at the site of discovery. The use of these test devices for detecting human blood at crime scenes is a common method. However, the IRT is specific not only for blood but also for differentiation between human and non-human soft tissue remains. In the following this method is discussed and validated by means of two forensic cases and several samples of various animals. Copyright © 2017 The Chartered Society of Forensic Sciences. Published by Elsevier B.V. All rights reserved.

  7. Evaluation of the Verigene® Blood Culture Nucleic Acid test for rapid identification of gram positive pathogens from positive blood cultures

    Directory of Open Access Journals (Sweden)

    Agnese Cellini

    2015-06-01

    Full Text Available Background. The rapid identification of the etiology and the evaluation of the antimicrobial susceptibility of the bacteria causing bacteremia is of outmost relevance to set up an adequate treatment of sepsis. In this study we evaluated the microarray based method, Verigene Gram-positive blood cultures (BC-GP nucleic acid test (Nanosphere Inc., Northbrook, IL, USA for the identification of Gram positive pathogens from positive blood cultures. The panel BC-GP is capable to identify 13 germs and 3 genes associated with antimicrobial resistance. Materials and Methods. In this study a total of 100 positive, non replicated and monomicrobic blood cultures have been evaluated. For testing on the Verigene platform using the BC-GP assay, 350 L of blood culture media from a positive the blood culture bottle.Results. A total of 100 positive blood cultures were tested by the Verigene BC-GP assay: out of these a total of 100 Gram-positive cocci were identified. The most frequent bacteria identified included staphylococci, streptococci and enterococci. Among staphylococci, Staphylococcus aureus accounted for 25% (15/60, with 38% of S. epidermidis 37% (23/60 and 37% (22/60 other CoNS. All the S. aureus isolates were correctly identified by BC-GP whereas in 2/45 cases (4% BC-GP misidentified CoNS. In the case of enterococci 7/10 were E. faecalis and 3 E. faecium, all of these were correctly identified.Conclusions. The overall agreement with the results obtained by standard procedure is quite elevated (88% and as a consequence the BC-GP panel could be used as a rapid diagnostic tool to give a faster response in the case of bacteremia associated with sepsis.

  8. Rapid and automatic chemical identification of the medicinal flower buds of Lonicera plants by the benchtop and hand-held Fourier transform infrared spectroscopy

    Science.gov (United States)

    Chen, Jianbo; Guo, Baolin; Yan, Rui; Sun, Suqin; Zhou, Qun

    2017-07-01

    With the utilization of the hand-held equipment, Fourier transform infrared (FT-IR) spectroscopy is a promising analytical technique to minimize the time cost for the chemical identification of herbal materials. This research examines the feasibility of the hand-held FT-IR spectrometer for the on-site testing of herbal materials, using Lonicerae Japonicae Flos (LJF) and Lonicerae Flos (LF) as examples. Correlation-based linear discriminant models for LJF and LF are established based on the benchtop and hand-held FT-IR instruments. The benchtop FT-IR models can exactly recognize all articles of LJF and LF. Although a few LF articles are misjudged at the sub-class level, the hand-held FT-IR models are able to exactly discriminate LJF and LF. As a direct and label-free analytical technique, FT-IR spectroscopy has great potential in the rapid and automatic chemical identification of herbal materials either in laboratories or in fields. This is helpful to prevent the spread and use of adulterated herbal materials in time.

  9. Single-cell pH imaging and detection for pH profiling and label-free rapid identification of cancer-cells.

    Science.gov (United States)

    Hou, Hui; Zhao, Yangyang; Li, Chuanping; Wang, Minmin; Xu, Xiaolong; Jin, Yongdong

    2017-05-11

    Single-cell pH-sensing and accurate detection and label-free fast identification of cancer-cells are two long-standing pursuits in cell and life science, as intracellular pH plays a crucial role in many cellular events and fates, while the latter is vital for early cancer theranostics. Numerous methods based on functionalized nanoparticles and fluorescence probes have been developed for cell pH-sensing, but are often hindered for single-cell studies by their main drawbacks of complicated probe preparation and labeling, low sensitivity and poor reproducibility. Here we report a simple and reliable method for single-cell pH imaging and sensing by innovative combined use of UV-Vis microspectroscopy and common pH indicators. Accurate and sensitive pH detection on single-cell or sub-cell level with good reproducibility is achieved by the method, which enables facile single-cell pH profiling and label-free rapid identification of cancer-cells (due to distinguishable intracellular pH levels) for early cancer diagnosis, and may open a new avenue for pH-related single-cell studies.

  10. Matrix-assisted laser desorption ionization time of flight mass spectrometry (MALDI-TOF MS) for rapid identification of micro-organisms in the routine clinical microbiology laboratory.

    Science.gov (United States)

    Wattal, C; Oberoi, J K; Goel, N; Raveendran, R; Khanna, S

    2017-05-01

    The study evaluates the utility of matrix-assisted laser desorption/ionisation time-of-flight mass spectrometry (MALDI-TOF MS) Vitek MS for identification of microorganisms in the routine clinical microbiology laboratory. From May 2013 to April 2014, microbial isolates recovered from various clinical samples were identified by Vitek MS. In case of failure to identify by Vitek MS, the isolate was identified using the Vitek 2 system (bioMerieux, France) and serotyping wherever applicable or otherwise by nucleic acid-mediated methods. All the moulds were identified by Lactophenol blue mounts, and mycobacterial isolates were identified by molecular identification systems including AccuProbe (bioMerieux, France) or GenoType Mycobacterium CM (Hain Lifescience, Germany). Out of the 12,003 isolates, the Vitek MS gave a good overall ID at the genus and or species level up to 97.7% for bacterial isolates, 92.8% for yeasts and 80% for filamentous fungi. Of the 26 mycobacteria tested, only 42.3% could be identified using the Saramis RUO (Research Use Only) database. VITEK MS could not identify 34 of the 35 yeast isolates identified as C. haemulonii by Vitek 2. Subsequently, 17 of these isolates were identified as Candida auris (not present in the Vitek MS database) by 18S rRNA sequencing. Using these strains, an in-house superspectrum of C. auris was created in the VITEK MS database. Use of MALDI-TOF MS allows a rapid identification of aerobic bacteria and yeasts in clinical practice. However, improved sample extraction protocols and database upgrades with inclusion of locally representative strains is required, especially for moulds.

  11. Microchip-based one step DNA extraction and real-time PCR in one chamber for rapid pathogen identification.

    Science.gov (United States)

    Lee, Jeong-Gun; Cheong, Kwang Ho; Huh, Nam; Kim, Suhyeon; Choi, Jeong-Woo; Ko, Christopher

    2006-07-01

    Optimal detection of a pathogen present in biological samples depends on the ability to extract DNA molecules rapidly and efficiently. In this paper, we report a novel method for efficient DNA extraction and subsequent real-time detection in a single microchip by combining laser irradiation and magnetic beads. By using a 808 nm laser and carboxyl-terminated magnetic beads, we demonstrate that a single pulse of 40 seconds lysed pathogens including E. coli and Gram-positive bacterial cells as well as the hepatitis B virus mixed with human serum. We further demonstrate that the real-time pathogen detection was performed with pre-mixed PCR reagents in a real-time PCR machine using the same microchip, after laser irradiation in a hand-held device equipped with a small laser diode. These results suggest that the new sample preparation method is well suited to be integrated into lab-on-a-chip application of the pathogen detection system.

  12. Rapid and Sensitive Identification of the Herbal Tea Ingredient Taraxacum formosanum Using Loop-Mediated Isothermal Amplification

    Directory of Open Access Journals (Sweden)

    Guan-Hua Lai

    2015-01-01

    Full Text Available Taraxacum formosanum (TF is a medicinal plant used as an important component of health drinks in Taiwan. In this study, a rapid, sensitive and specific loop-mediated isothermal amplification (LAMP assay for authenticating TF was established. A set of four specific LAMP primers was designed based on the nucleotide sequence of the internal transcribed spacer 2 (ITS2 nuclear ribosomal DNA (nrDNA of TF. LAMP amplicons were successfully amplified and detected when purified genomic DNA of TF was added in the LAMP reaction under isothermal condition (65 °C within 45 min. These specific LAMP primers have high specificity and can accurately discriminate Taraxacum formosanum from other adulterant plants; 1 pg of genomic DNA was determined to be the detection limit of the LAMP assay. In conclusion, using this novel approach, TF and its misused plant samples obtained from herbal tea markets were easily identified and discriminated by LAMP assay for quality control.

  13. Rapid identification of molecular changes in tulsi (Ocimum sanctum Linn) upon ageing using leaf spray ionization mass spectrometry.

    Science.gov (United States)

    Sarkar, Depanjan; Srimany, Amitava; Pradeep, T

    2012-10-07

    Tulsi or Holy Basil (Ocimum sanctum Linn) is a medicinally important plant. Ursolic acid (UA) and oleanolic acid (OA) are among its major constituents which account for many medicinal activities of the plant. In the present work, we deployed a new ambient ionization method, leaf spray ionization, for rapid detection of UA, OA and their oxidation products from tulsi leaves. Tandem electrospray ionization mass spectrometry (ESI-MS) has been performed on tulsi leaf extracts in methanol to establish the identity of the compounds. We probed changes occurring in the relative amounts of the parent compounds (UA and OA) with their oxidized products and the latter show an increasing trend upon ageing. The findings are verified by ESI-MS analysis of tulsi leaf extracts, which shows the same trend proving the reliability of the leaf spray method.

  14. Accurate, rapid identification of dislocation lines in coherent diffractive imaging via a min-max optimization formulation

    Science.gov (United States)

    Ulvestad, A.; Menickelly, M.; Wild, S. M.

    2018-01-01

    Defects such as dislocations impact materials properties and their response during external stimuli. Imaging these defects in their native operating conditions to establish the structure-function relationship and, ultimately, to improve performance via defect engineering has remained a considerable challenge for both electron-based and x-ray-based imaging techniques. While Bragg coherent x-ray diffractive imaging (BCDI) is successful in many cases, nuances in identifying the dislocations has left manual identification as the preferred method. Derivative-based methods are also used, but they can be inaccurate and are computationally inefficient. Here we demonstrate a derivative-free method that is both more accurate and more computationally efficient than either derivative- or human-based methods for identifying 3D dislocation lines in nanocrystal images produced by BCDI. We formulate the problem as a min-max optimization problem and show exceptional accuracy for experimental images. We demonstrate a 227x speedup for a typical experimental dataset with higher accuracy over current methods. We discuss the possibility of using this algorithm as part of a sparsity-based phase retrieval process. We also provide MATLAB code for use by other researchers.

  15. Metabolomics beyond spectroscopic databases: a combined MS/NMR strategy for the rapid identification of new metabolites in complex mixtures.

    Science.gov (United States)

    Bingol, Kerem; Bruschweiler-Li, Lei; Yu, Cao; Somogyi, Arpad; Zhang, Fengli; Brüschweiler, Rafael

    2015-04-07

    A novel strategy is introduced that combines high-resolution mass spectrometry (MS) with NMR for the identification of unknown components in complex metabolite mixtures encountered in metabolomics. The approach first identifies the chemical formulas of the mixture components from accurate masses by MS and then generates all feasible structures (structural manifold) that are consistent with these chemical formulas. Next, NMR spectra of each member of the structural manifold are predicted and compared with the experimental NMR spectra in order to identify the molecular structures that match the information obtained from both the MS and NMR techniques. This combined MS/NMR approach was applied to Escherichia coli extract, where the approach correctly identified a wide range of different types of metabolites, including amino acids, nucleic acids, polyamines, nucleosides, and carbohydrate conjugates. This makes this approach, which is termed SUMMIT MS/NMR, well suited for high-throughput applications for the discovery of new metabolites in biological and biomedical mixtures, overcoming the need of experimental MS and NMR metabolite databases.

  16. Use of a conformational switching aptamer for rapid and specific ex vivo identification of central nervous system lymphoma in a xenograft model.

    Directory of Open Access Journals (Sweden)

    Joseph F Georges

    Full Text Available Improved tools for providing specific intraoperative diagnoses could improve patient care. In neurosurgery, intraoperatively differentiating non-operative lesions such as CNS B-cell lymphoma from operative lesions can be challenging, often necessitating immunohistochemical (IHC procedures which require up to 24-48 hours. Here, we evaluate the feasibility of generating rapid ex vivo specific labeling using a novel lymphoma-specific fluorescent switchable aptamer. Our B-cell lymphoma-specific switchable aptamer produced only low-level fluorescence in its unbound conformation and generated an 8-fold increase in fluorescence once bound to its target on CD20-positive lymphoma cells. The aptamer demonstrated strong binding to B-cell lymphoma cells within 15 minutes of incubation as observed by flow cytometry. We applied the switchable aptamer to ex vivo xenograft tissue harboring B-cell lymphoma and astrocytoma, and within one hour specific visual identification of lymphoma was routinely possible. In this proof-of-concept study in human cell culture and orthotopic xenografts, we conclude that a fluorescent switchable aptamer can provide rapid and specific labeling of B-cell lymphoma, and that developing aptamer-based labeling approaches could simplify tissue staining and drastically reduce time to histopathological diagnoses compared with IHC-based methods. We propose that switchable aptamers could enhance expeditious, accurate intraoperative decision-making.

  17. Development of a Loop-Mediated Isothermal Amplification Assay for Rapid and Specific Identification of ACT Producing Alternaria alternata, the Agent of Brown Spot Disease in Tangerine.

    Science.gov (United States)

    Moghimi, Hamid; Moradi, Amir; Hamedi, Javad; Basiri, Mina

    2016-03-01

    Rapid, accurate, and easy identification of pathogenic agents has always been important in medicine, veterinary, and agriculture. The brown spot infection is among the most common diseases in tangerine caused by Alternaria alternata. Due to the existence of seven various pathotypes of A. alternata species, it is challenging and time consuming to detect a pathotype responsible for citrus brown spot. In this study, we were seeking a rapid and specific approach to identify the tangerine pathotype within the A. alternata-pathogenic species, using the loop-mediated isothermal amplification (LAMP) method and actts2 gene as a marker molecule. Nine pathogenic samples were obtained from the region of Ramsar, Iran, and certified as A. alternata-pathogenic isolates. Specific primers were designed for regions coding for Alternaria citri toxin (ACT), and the PCR and LAMP reactions were performed. Our data showed that the primers designed for the tangerine pathotype of A. alternata were specific, and in both reactions, positive results were only observed in desired pathotypes. In the other pathotypes of this species as well as other standard fungal samples as negative controls, no positive result was observed. Therefore, our results suggest the possibility to detect the tangerine-specific A. alternata pathotype from other related species with a high accuracy and in early stages of the disease.

  18. Rapid Genotyping of the Human Renin (REN Gene by the LightCycler® Instrument: Identification of Unexpected Nucleotide Substitutions within the Selected Hybridization Probe Area

    Directory of Open Access Journals (Sweden)

    Line Wee

    2010-01-01

    Full Text Available Preeclampsia is a serious disorder affecting nearly 3% of all in the Western world. It is associated with hypertension and proteinuria, and several lines of evidence suggest that the renin-angiotensin system (RAS may be involved in the development of hypertension at different stages of a preeclamptic pregnancy. In this study, we developed rapid genotyping assays on the LightCycler® instrument to allow the detection of genetic variants in the renin gene (REN that may predispose to preeclampsia. The method is based on real-time PCR and allele-specific hybridization probes, followed by fluorescent melting curve analysis to expose a change in melting temperature (Tm. Ninety-two mother-father-child triads (n=276 from preeclamptic pregnancies were genotyped for three haplotype-tagging single nucleotide polymorphisms (htSNPs in REN. All three htSNPs (rs5705, rs1464816 and rs3795575 were successfully genotyped. Furthermore, two unexpected nucleotide substitutions (rs11571084 and rs61757041 were identified within the selected hybridization probe area of rs1464816 and rs3795575 due to aberrant melting peaks. In conclusion, genotyping on the LightCycler® instrument proved to be rapid and highly reproducible. The ability to uncover additional nucleotide substitutions is particularly important in that it allows the identification of potentially etiological variants that might otherwise be overlooked by other genotyping methods.

  19. Rapid detection and identification of Wuchereria bancrofti, Brugia malayi, B. pahangi, and Dirofilaria immitis in mosquito vectors and blood samples by high resolution melting real-time PCR.

    Science.gov (United States)

    Thanchomnang, Tongjit; Intapan, Pewpan M; Tantrawatpan, Chairat; Lulitanond, Viraphong; Chungpivat, Sudchit; Taweethavonsawat, Piyanan; Kaewkong, Worasak; Sanpool, Oranuch; Janwan, Penchom; Choochote, Wej; Maleewong, Wanchai

    2013-12-01

    A simple, rapid, and high-throughput method for detection and identification of Wuchereria bancrofti, Brugia malayi, Brugia pahangi, and Dirofilaria immitis in mosquito vectors and blood samples was developed using a real-time PCR combined with high-resolution melting (HRM) analysis. Amplicons of the 4 filarial species were generated from 5S rRNA and spliced leader sequences by the real-time PCR and their melting temperatures were determined by the HRM method. Melting of amplicons from W. bancrofti, B. malayi, D. immitis, and B. pahangi peaked at 81.5±0.2℃, 79.0±0.3℃, 76.8±0.1℃, and 79.9±0.1℃, respectively. This assay is relatively cheap since it does not require synthesis of hybridization probes. Its sensitivity and specificity were 100%. It is a rapid and technically simple approach, and an important tool for population surveys as well as molecular xenomonitoring of parasites in vectors.

  20. Antibiotic treatment algorithm development based on a microarray nucleic acid assay for rapid bacterial identification and resistance determination from positive blood cultures.

    Science.gov (United States)

    Rödel, Jürgen; Karrasch, Matthias; Edel, Birgit; Stoll, Sylvia; Bohnert, Jürgen; Löffler, Bettina; Saupe, Angela; Pfister, Wolfgang

    2016-03-01

    Rapid diagnosis of bloodstream infections remains a challenge for the early targeting of an antibiotic therapy in sepsis patients. In recent studies, the reliability of the Nanosphere Verigene Gram-positive and Gram-negative blood culture (BC-GP and BC-GN) assays for the rapid identification of bacteria and resistance genes directly from positive BCs has been demonstrated. In this work, we have developed a model to define treatment recommendations by combining Verigene test results with knowledge on local antibiotic resistance patterns of bacterial pathogens. The data of 275 positive BCs were analyzed. Two hundred sixty-three isolates (95.6%) were included in the Verigene assay panels, and 257 isolates (93.5%) were correctly identified. The agreement of the detection of resistance genes with subsequent phenotypic susceptibility testing was 100%. The hospital antibiogram was used to develop a treatment algorithm on the basis of Verigene results that may contribute to a faster patient management. Copyright © 2016 Elsevier Inc. All rights reserved.

  1. Rapid Identification of the Mycobacterium tuberculosis Complex by Combining the ESAT-6/CFP-10 Immunochromatographic Assay and Smear Morphology▿

    Science.gov (United States)

    Shen, Gwan-Han; Chiou, Chien-Shun; Hu, Shiau-Ting; Wu, Kun-Ming; Chen, Jiann-Hwa

    2011-01-01

    Early secretory antigen 6 (ESAT-6) and cell filtrate protein 10 (CFP-10) are two antigens secreted as a complex by the replicating Mycobacterium tuberculosis complex (MTC). Recently, an immunochromatographic assay (ICA) using a monoclonal antibody against the ESAT-6/CFP-10 complex was developed for the purpose of MTC detection. In this study, the efficacy of the assay was tested with 603 BACTEC cultures that were incubated for 3 additional days after positive signals appeared in the BACTEC MGIT 960 system. Bacterial isolates were recovered from these 603 BACTEC cultures, and 332 MTC isolates, 270 nontuberculosis mycobacterial isolates, and 1 Nocardia isolate were identified by using standard biochemical assays. The ESAT-6/CFP-10 assay detected 322 MTC cultures, resulting in a sensitivity of 97% and a specificity of 97.4%. To reduce the false-negative rate and improve the sensitivity, either serpentine cording in an acid-fast bacillus stain of the cultural smear, the ESAT-6/CFP-10 assay, or a combination of both was used for MTC detection. The sensitivity was then increased to 99.1%, and the negative predictive value increased to 98.9%, but the specificity decreased to 94.8% and the positive predictive value decreased to 95.9%. However, a combination of serpentine cording in cultural smears and the positivity of the ICA resulted in the specificity and positive predictive values of 100%. Therefore, BACTEC cultures with both serpentine cording and positivity of the ESAT-6/CFP-10 assay could be reported to contain MTC directly. The ESAT-6/CFP-10 assay may be an alternative of the Capilia assay (MPB64-ICA) as a convenient and cost-effective method for identification of MTC in culture. PMID:21159936

  2. Rapid identification of the Mycobacterium tuberculosis complex by combining the ESAT-6/CFP-10 immunochromatographic assay and smear morphology.

    Science.gov (United States)

    Shen, Gwan-Han; Chiou, Chien-Shun; Hu, Shiau-Ting; Wu, Kun-Ming; Chen, Jiann-Hwa

    2011-03-01

    Early secretory antigen 6 (ESAT-6) and cell filtrate protein 10 (CFP-10) are two antigens secreted as a complex by the replicating Mycobacterium tuberculosis complex (MTC). Recently, an immunochromatographic assay (ICA) using a monoclonal antibody against the ESAT-6/CFP-10 complex was developed for the purpose of MTC detection. In this study, the efficacy of the assay was tested with 603 BACTEC cultures that were incubated for 3 additional days after positive signals appeared in the BACTEC MGIT 960 system. Bacterial isolates were recovered from these 603 BACTEC cultures, and 332 MTC isolates, 270 nontuberculosis mycobacterial isolates, and 1 Nocardia isolate were identified by using standard biochemical assays. The ESAT-6/CFP-10 assay detected 322 MTC cultures, resulting in a sensitivity of 97% and a specificity of 97.4%. To reduce the false-negative rate and improve the sensitivity, either serpentine cording in an acid-fast bacillus stain of the cultural smear, the ESAT-6/CFP-10 assay, or a combination of both was used for MTC detection. The sensitivity was then increased to 99.1%, and the negative predictive value increased to 98.9%, but the specificity decreased to 94.8% and the positive predictive value decreased to 95.9%. However, a combination of serpentine cording in cultural smears and the positivity of the ICA resulted in the specificity and positive predictive values of 100%. Therefore, BACTEC cultures with both serpentine cording and positivity of the ESAT-6/CFP-10 assay could be reported to contain MTC directly. The ESAT-6/CFP-10 assay may be an alternative of the Capilia assay (MPB64-ICA) as a convenient and cost-effective method for identification of MTC in culture.

  3. Using automated medical records for rapid identification of illness syndromes (syndromic surveillance: the example of lower respiratory infection

    Directory of Open Access Journals (Sweden)

    Dashevsky Inna

    2001-10-01

    Full Text Available Abstract Background Gaps in disease surveillance capacity, particularly for emerging infections and bioterrorist attack, highlight a need for efficient, real time identification of diseases. Methods We studied automated records from 1996 through 1999 of approximately 250,000 health plan members in greater Boston. Results We identified 152,435 lower respiratory infection illness visits, comprising 106,670 episodes during 1,143,208 person-years. Three diagnoses, cough (ICD9CM 786.2, pneumonia not otherwise specified (ICD9CM 486 and acute bronchitis (ICD9CM 466.0 accounted for 91% of these visits, with expected age and sex distributions. Variation of weekly occurrences corresponded closely to national pneumonia and influenza mortality data. There was substantial variation in geographic location of the cases. Conclusion This information complements existing surveillance programs by assessing the large majority of episodes of illness for which no etiologic agents are identified. Additional advantages include: a sensitivity, uniformity and efficiency, since detection of events does not depend on clinicians' to actively report diagnoses, b timeliness, the data are available within a day of the clinical event; and c ease of integration into automated surveillance systems. These features facilitate early detection of conditions of public health importance, including regularly occurring events like seasonal respiratory illness, as well as unusual occurrences, such as a bioterrorist attack that first manifests as respiratory symptoms. These methods should also be applicable to other infectious and non-infectious conditions. Knowledge of disease patterns in real time may also help clinicians to manage patients, and assist health plan administrators in allocating resources efficiently.

  4. Rapid identification of siderophores by combined thin-layer chromatography/matrix-assisted laser desorption/ionization mass spectrometry.

    Science.gov (United States)

    Hayen, Heiko; Volmer, Dietrich A

    2005-01-01

    The investigation of a combined thin-layer chromatography/matrix-assisted laser desorption/ionization mass spectrometry (TLC/MALDI-MS) method for the analysis of siderophores from microbial samples is described. The investigated siderophores were enterobactin, ferrioxamine B, ferrichrome, ferrirhodin, rhodotorulic acid and coprogen. Solid-phase extraction was employed to recover the siderophores from the microbial samples. After visualization of the spots via spraying with ferric chloride or chrome azurol sulfonate assay solution, the MALDI matrix was applied to the gel surface. Several TLC/MALDI experimental parameters were optimized, such as type and concentration of MALDI matrix, as well as the type and composition of solvent to facilitate analyte transport from the inside of the TLC gel to the surface. The impact of these parameters on sensitivity, precision and ion formation of the various siderophores was studied. The detection limits for the investigated siderophores were in the range 1-4 pmol. These values were about 4-24 times higher than the detection limits obtained directly from stainless steel MALDI targets. The differences were most likely due to incomplete transport of the 'trapped' analyte molecules from the deeper layers of the TLC gel to the surface and into the matrix layer. In addition, chromatographic band broadening spread the analyte further in TLC as compared with the steel plates, resulting in less analyte per surface area. The identification of the siderophores was aided by concurrently applying a Ga(III) nitrate solution to the TLC plate during the visualization step. The resulting formation of Ga(III) complexes lead to distinctive (69)Ga/(71)Ga isotope patterns in the mass spectra. The versatility of the TLC/MALDI-MS assay was demonstrated by using it to analyze siderophores in a Pseudomonas aeruginosa sample. An iron-binding compound was identified in the sample, namely pyochelin (2-(2-o-hydroxyphenyl-2-thiazolin-4-yl)-3

  5. Rapid identification of Mycobacterium tuberculosis infection by a new array format-based surface plasmon resonance method

    Science.gov (United States)

    Hsieh, Shang-Chen; Chang, Chia-Chen; Lu, Chia-Chen; Wei, Chia-Fong; Lin, Chuan-Sheng; Lai, Hsin-Chih; Lin, Chii-Wann

    2012-03-01

    Tubercle bacillus [TB] is one of the most important chronic infectious diseases that cause millions of deaths annually. While conventional smear microscopy and culture methods are widely used for diagnosis of TB, the former is insensitive, and the latter takes up to 6 to 8 weeks to provide a result, limiting the value of these methods in aiding diagnosis and intermediate decisions on treatment. Therefore, a rapid detection method is essential for the diagnosis, prognosis assessment, and recurrence monitoring. A new surface plasmon resonance [SPR] biosensor based on an array format, which allowed immobilizing nine TB antigens onto the sensor chip, was constructed. Simultaneous determination of multiple TB antibodies in serum had been accomplished with this array-based SPR system. The results were compared with enzyme-linked immunosorbent assay, a conventional immunological method. Array-based SPR showed more advantages in providing label-free and real-time detection. Additionally, the high sensitivity and specificity for the detection of TB infection showed its potential for future development of biosensor arrays for TB diagnosis.

  6. Rapid presumptive identification of the Mycobacterium tuberculosis-bovis complex by radiometric determination of heat stable urease

    Energy Technology Data Exchange (ETDEWEB)

    Gandy, J.H.; Pruden, E.L.; Cox, F.R.

    1983-12-01

    Simple and rapid Bactec methodologies for the determination of neat (unaltered) and heat stable urease activity of mycobacteria are presented. Clinical isolates (63) and stock cultures (32)--consisting of: M. tuberculosis (19), M. bovis (5), M. kansasii (15), M. marinum (4), M. simiae (3), M. scrofulaceum (16), M. gordonae (6), M. szulgai (6), M. flavescens (1), M. gastri (1), M. intracellulare (6), M. fortuitum-chelonei complex (12), and M. smegmatis (1)--were tested for neat urease activity by Bactec radiometry. Mycobacterial isolates (50-100 mg wet weight) were incubated at 35 degrees C for 30 minutes with microCi14C-urea. Urease-positive mycobacteria gave Bactec growth index (GI) values greater than 100 units, whereas urease-negative species gave values less than 10 GI units. Eighty-three isolates possessing neat urease activity were heated at 80 degrees C for 30 minutes followed by incubation at 35 degrees C for 30 minutes with 1 microCi14C-urea. Mycobacterium tuberculosis-bovis complex demonstrated heat-stable urease activity (GI more than 130 units) and could be distinguished from mycobacteria other than tuberculosis (MOTT), which gave GI values equal to or less than 40 units.

  7. Rapid antimicrobial susceptibility test for identification of new therapeutics and drug combinations against multidrug-resistant bacteria

    Science.gov (United States)

    Sun, Wei; Weingarten, Rebecca A; Xu, Miao; Southall, Noel; Dai, Sheng; Shinn, Paul; Sanderson, Philip E; Williamson, Peter R; Frank, Karen M; Zheng, Wei

    2016-01-01

    Current antimicrobial susceptibility testing has limited screening capability for identifying empirical antibiotic combinations to treat severe bacterial infections with multidrug-resistant (MDR) organisms. We developed a new antimicrobial susceptibility assay using automated ultra-high-throughput screen technology in combination with a simple bacterial growth assay. A rapid screening of 5170 approved drugs and other compounds identified 25 compounds with activities against MDR Klebsiella pneumoniae. To further improve the efficacy and reduce the effective drug concentrations, we applied a targeted drug combination approach that integrates drugs' clinical antimicrobial susceptibility breakpoints, achievable plasma concentrations, clinical toxicities and mechanisms of action to identify optimal drug combinations. Three sets of three-drug combinations were identified with broad-spectrum activities against 10 MDR clinical isolates including K. pneumoniae, Acinetobacter baumannii, Pseudomonas aeruginosa, Citrobacter freundii, Enterobacter cloacae and Escherichia coli. Colistin–auranofin–ceftazidime and colistin–auranofin–rifabutin suppressed >80% growth of all 10 MDR strains; while rifabutin–colistin–imipenem inhibited >75% of these strains except two Acinetobacter baumannii isolates. The results demonstrate this new assay has potential as a real-time method to identify new drugs and effective drug combinations to combat severe clinical infections with MDR organisms. PMID:27826141

  8. A new method for rapid identification of ansamycin compounds by inactivating KLM gene clusters in potential ansamycin-producing actinomyces.

    Science.gov (United States)

    Wang, G; Zhang, H; Sun, G; Wu, L; Zhang, J; Wang, Y

    2012-02-01

    In this study, we explored the possibility of construction of a 'universal targeting vector' by Red/ET recombination to inactivate L gene encoding 3-amino-5-hydroxybenzoic acid (AHBA)-oxidoreductase in AHBA biosynthetic gene cluster to facilitate the detection of ansamycins production in actinomycetes. Based on the conserved regions of linked AHBA synthase (K), oxidoreductase (L) and phosphatase (M) gene clusters, degenerate primers were designed and PCR was performed to detect KLM gene clusters within 33 AHBA synthase gene-positive actinomycetes strains. Among them, 22 KLM gene cluster-positive strains were identified. A 'universal targeting vector' was further constructed using the 50-nt homologous sequences chosen from four strains internal L gene in KLM gene clusters through Red/ET recombination. The L gene from nine of the KLM gene cluster-positive actinomycetes strains was inactivated by insertion of a kanamycin (Km) resistance marker into its internal region from the 'universal targeting vector'. By comparison of the metabolites produced in parent strains with those in L gene-inactivated mutants, we demonstrated the possible ansamycins production produced by these strains. One strain (4089) was proved to be a geldanamycin producer. Three strains (3-20, 7-32 and 8-32) were identified as potential triene-ansamycins producers. Another strain (3-27) was possible to be a streptovaricin C producer. Strains 24-100 and 4-124 might be served as ansamitocin-like producers. The results confirmed the feasibility that a 'universal targeting vector' could be constructed through Red/ET recombination using the conserved regions of KLM gene clusters to detect ansamycins production in actinomycetes. The 'universal targeting vector' provides a rapid approach in certain degree to detect the potential ansamycin producers from the 22 KLM gene cluster-positive actinomycetes strains. © 2011 The Authors. Journal of Applied Microbiology © 2011 The Society for Applied Microbiology.

  9. A new real-time PCR assay for rapid identification of the S. aureus/MRSA strains

    Directory of Open Access Journals (Sweden)

    Ivan Manga

    2013-01-01

    Full Text Available The Methicillin-resistant Staphylococcus aureus (MRSA with the livestock-associated MRSA (LA-MRSA are of great interest to scientists and general public. The aim of our study was to present a new more rapid and reliable diagnostic method working on the RT-PCR platform applicable for monitoring of MRSA/S. aureus. The parallel testing of the S. aureus specific nuc gene sequence and the mecA gene sequence was utilised for this purpose. A collection of ten S. aureus/MRSA reference strains, fifteen genetically related non S. aureus reference strains and fifty-six environmental samples was employed for estimation of the assay performance and parameters. The environmental samples acquired in the Czech livestock farms were represented with the livestock and human nasal mucosae or skin swabs, the slaughter meat swabs and were chosen preferentially from individuals with previously confi rmed or suspected positive MRSA/S. aureus cases. The classic selective cultivation approach with the biochemical test and agar disk diffusion test was accepted as reference diagnostic method. As there were no culture positive samples that were negative using RT-PCR, our method featured with 100% sensitivity in comparison to reference method. The limit of detection allowed to identify from tens to hundreds copies of S. aureus/MRSA genome. Further, the RT-PCR assay featured with 100% inclusivity and 95% exclusivity at Cq value below 30. These parameters suggested on powerful and reliable diagnostic method with real potential of practical utilisation. We consider our method as ideal for testing of individual suspected colonies, when the results can be acquired in less than 1.5 hour.

  10. A novel rapid DNA microarray assay enables identification of 37 Mycoplasma species and highlights multiple Mycoplasma infections.

    Directory of Open Access Journals (Sweden)

    Christiane Schnee

    Full Text Available Mycoplasmas comprise a conglomerate of pathogens and commensals occurring in humans and animals. The genus Mycoplasma alone contains more than 120 species at present, and new members are continuously being discovered. Therefore, it seems promising to use a single highly parallel detection assay rather than develop separate tests for each individual species. In this study, we have designed a DNA microarray carrying 70 oligonucleotide probes derived from the 23S rRNA gene and 86 probes from the tuf gene target regions. Following a PCR amplification and biotinylation step, hybridization on the array was shown to specifically identify 31 Mycoplasma spp., as well as 3 Acholeplasma spp. and 3 Ureaplasma spp. Members of the Mycoplasma mycoides cluster can be recognized at subgroup level. This procedure enables parallel detection of Mollicutes spp. occurring in humans, animals or cell culture, from mono- and multiple infections, in a single run. The main advantages of the microarray assay include ease of operation, rapidity, high information content, and affordability. The new test's analytical sensitivity is equivalent to that of real-time PCR and allows examination of field samples without the need for culture. When 60 field samples from ruminants and birds previously analyzed by denaturing-gradient gel electrophoresis (DGGE were tested by the microarray assay both tests identified the same agent in 98.3% of the cases. Notably, microarray testing revealed an unexpectedly high proportion (35% of multiple mycoplasma infections, i.e., substantially more than DGGE (15%. Two of the samples were found to contain four different Mycoplasma spp. This phenomenon deserves more attention, particularly its implications for epidemiology and treatment.

  11. Novel, Rapid Identification, and Quantification of Adulterants in Extra Virgin Olive Oil Using Near-Infrared Spectroscopy and Chemometrics.

    Science.gov (United States)

    Azizian, Hormoz; Mossoba, Magdi M; Fardin-Kia, Ali Reza; Delmonte, Pierluigi; Karunathilaka, Sanjeewa R; Kramer, John K G

    2015-07-01

    A new, rapid Fourier transform near infrared (FT-NIR) spectroscopic procedure is described to screen for the authenticity of extra virgin olive oils (EVOO) and to determine the kind and amount of an adulterant in EVOO. To screen EVOO, a partial least squares (PLS1) calibration model was developed to estimate a newly created FT-NIR index based mainly on the relative intensities of two unique carbonyl overtone absorptions in the FT-NIR spectra of EVOO and other mixtures attributed to volatile (5280 cm(-1)) and non-volatile (5180 cm(-1)) components. Spectra were also used to predict the fatty acid (FA) composition of EVOO or samples spiked with an adulterant using previously developed PLS1 calibration models. Some adulterated mixtures could be identified provided the FA profile was sufficiently different from those of EVOO. To identify the type and determine the quantity of an adulterant, gravimetric mixtures were prepared by spiking EVOO with different concentrations of each adulterant. Based on FT-NIR spectra, four PLS1 calibration models were developed for four specific groups of adulterants, each with a characteristic FA composition. Using these different PLS1 calibration models for prediction, plots of predicted vs. gravimetric concentrations of an adulterant in EVOO yielded linear regression functions with four unique sets of slopes, one for each group of adulterants. Four corresponding slope rules were defined that allowed for the determination of the nature and concentration of an adulterant in EVOO products by applying these four calibration models. The standard addition technique was used for confirmation.

  12. Primate genotyping via high resolution melt analysis: rapid and reliable identification of color vision status in wild lemurs.

    Science.gov (United States)

    Jacobs, Rachel L; Spriggs, Amanda N; MacFie, Tammie S; Baden, Andrea L; Irwin, Mitchell T; Wright, Patricia C; Louis, Edward E; Lawler, Richard R; Mundy, Nicholas I; Bradley, Brenda J

    2016-10-01

    Analyses of genetic polymorphisms can aid our understanding of intra- and interspecific variation in primate sociality, ecology, and behavior. Studies of primate opsin genes are prime examples of this, as single nucleotide variants (SNVs) in the X-linked opsin gene underlie variation in color vision. For primate species with polymorphic trichromacy, genotyping opsin SNVs can generally indicate whether individual primates are red-green color-blind (denoted homozygous M or homozygous L) or have full trichromatic color vision (heterozygous ML). Given the potential influence of color vision on behavior and fitness, characterizing the color vision status of study subjects is becoming commonplace for many primate field projects. Such studies traditionally involve a multi-step sequencing-based method that can be costly and time-consuming. Here we present a new reliable, rapid, and relatively inexpensive method for characterizing color vision in primate populations using high resolution melt analysis (HRMA). Using lemurs as a case study, we characterized variation at exons 3 and/or 5 of the X-linked opsin gene for 87 individuals representing nine species. We scored opsin genotypes and color vision status using both traditional sequencing-based methods as well as our novel melting-curve based HRMA protocol. For each species, the melting curves of varying genotypes (homozygous M, homozygous L, heterozygous ML) differed in melting temperature and/or shape. Melting curves for each sample were consistent across replicates, and genotype-specific melting curves were consistent across DNA sources (blood vs. feces). We show that opsin genotypes can be quickly and reliably scored using HRMA once lab-specific reference curves have been developed based on known genotypes. Although the protocol presented here focuses on genotyping lemur opsin loci, we also consider the larger potential for applying this approach to various types of genetic studies of primate populations.

  13. Stepwise discriminant function analysis for rapid identification of acute promyelocytic leukemia from acute myeloid leukemia with multiparameter flow cytometry.

    Science.gov (United States)

    Chen, Zhanguo; Li, Yan; Tong, Yongqing; Gao, Qingping; Mao, Xiaolu; Zhang, Wenjing; Xia, Zunen; Fu, Chaohong

    2016-03-01

    Diagnosis of acute promyelocytic leukemia (APL) has been accelerated by multiparameter flow cytometry (MFC). However, diagnostic interpretation of MFC readouts for APL depends on individual experience and knowledge, which inevitably increases the risk of arbitrariness. We appraised the feasibility of using stepwise discriminant function analysis (SDFA) based on MFC to optimize the minimal variables needed to distinguish APL from other acute myeloid leukemia (AML) without complicated data interpretation. Samples from 327 patients with APL (n = 51) and non-APL AML (n = 276) were randomly allocated into training (243 AML) and test sets (84 AML) for SDFA. The discriminant functions from SDFA were examined by correct classification, and the final variables were validated by differential expression. Finally, additional 20 samples from patients with atypical APL and AML confusable with APL were also identified by SDFA method and morphological analysis. The weighed discriminant function reveals seven differentially expressed variables (CD2/CD9/CD11b/CD13/CD34/HLA-DR/CD117), which predict a molecular result for APL characterization with an accuracy that approaches 99% (99.6 and 98.8% for AML samples in training and test sets, respectively). Furthermore, the SDFA outperformed either single variable analysis or the more limited 3-component analysis (CD34/CD117/HLA-DR) via separate SDFA, and was also superior to morphological analysis in terms of diagnostic efficacy. The established SDFA based on MFC with seven variables can precisely and rapidly differentiate APL and non-APL AML, which may contribute to the urgent initiation of all-trans-retinoic acid-based APL therapy.

  14. Rapid screening and identification of chemical hazards in surface and drinking water using high resolution mass spectrometry and a case-control filter.

    Science.gov (United States)

    Kaserzon, Sarit L; Heffernan, Amy L; Thompson, Kristie; Mueller, Jochen F; Gomez Ramos, Maria Jose

    2017-09-01

    Access to clean, safe drinking water poses a serious challenge to regulators, and requires analytical strategies capable of rapid screening and identification of potentially hazardous chemicals, specifically in situations when threats to water quality or security require rapid investigations and potential response. This study describes a fast and efficient chemical hazard screening strategy for characterising trace levels of polar organic contaminants in water matrices, based on liquid chromatography high resolution mass spectrometry with post-acquisition 'case-control' data processing. This method allowed for a rapid response time of less than 24 h for the screening of target, suspect and non-target unknown chemicals via direct injection analysis, and a second, more sensitive analysis option requiring sample pre-concentration. The method was validated by fortifying samples with a range of pesticides, pharmaceuticals and personal care products (n = 46); with >90% of target compounds positively screened in samples at 1 ng mL-1, and 46% at 0.1 ng mL-1 when analysed via direct injection. To simulate a contamination event samples were fortified with compounds not present in the commercial library (designated 'non-target compounds'; fipronil and fenitrothion), tentatively identified at 0.2 and 1 ng mL-1, respectively; and a compound not included in any known commercial library or public database (designated 'unknown' compounds; 8Cl- perfluorooctanesulfonic acid), at 0.8 ng mL-1. The method was applied to two 'real-case' scenarios: (1) the assessment of drinking water safety during a high-profile event in Brisbane, Australia; and (2) to screen treated, re-circulated drinking water and pre-treated (raw) water. The validated workflow was effective for rapid prioritisation and screening of suspect and non-target potential hazards at trace levels, and could be applied to a wide range of matrices and investigations where comparison of organic contaminants between

  15. EVALUATION OF A RAPID SCREENING ASSAY FOR BACTERIAL IDENTIFICATION (DOT-ELISA IN FECAL SAMPLES FROM CHILDREN

    Directory of Open Access Journals (Sweden)

    Etelvina BOCCATTO

    1997-01-01

    Full Text Available With the objective of standardizing a Dot Enzyme-Linked Immunosorbent Assay (Dot-ELISA to detect antigens of fecal bacterial enteropathogens, 250 children, aged under 36 months and of both sexes, were studied; of which 162 had acute gastroenteritis. The efficacy of a rapid screening assay for bacterial enteropathogens (enteropathogenic Escherichia coli "EPEC", enteroinvasive Escherichia coli "EIEC", Salmonella spp. and Shigella spp. was evaluated. The fecal samples were also submitted to a traditional method of stool culture for comparison. The concordance index between the two techniques, calculated using the Kappa (k index for the above mentioned bacterial strains was 0.8859, 0.9055, 0.7932 and 0.7829 respectively. These values express an almost perfect degree of concordance for the first two and substantial concordance for the latter two, thus enabling this technique to be applied in the early diagnosis of diarrhea in infants. With a view to increasing the sensitivity and specificity of this immunological test, a study was made of the antigenic preparations obtained from two types of treatment: 1 deproteinization by heating; 2 precipitation and concentration of the lipopolysaccharide antigen (LPS using an ethanol-acetone solution, which was then heated in the presence of sodium EDTACom o objetivo de padronizar um Dot Enzyme-Linked Immunosorbent Assay (Dot-ELISA para a detecção de antígenos de enteropatógenos bacterianos fecais, estudaram-se 250 crianças, abaixo de 36 meses de idade, de ambos os sexos, 162 portadoras de gastroenterite aguda. Avaliou-se a eficácia de um teste rápido para bactérias enteropatógenas (Escherichia coli enteropatogênica "EPEC", Escherichia coli enteroinvasora" EIEC", Salmonella spp. e Shigella spp.. As amostras fecais foram também submetidas à metodologia tradicional de coprocultura para comparação. Os índices de concordância entre as 2 técnicas, calculado através do índice Kappa (k para as cepas

  16. Establishment of a simple and rapid identification method for Listeria spp. by using high-resolution melting analysis, and its application in food industry.

    Science.gov (United States)

    Ohshima, Chihiro; Takahashi, Hajime; Phraephaisarn, Chirapiphat; Vesaratchavest, Mongkol; Keeratipibul, Suwimon; Kuda, Takashi; Kimura, Bon

    2014-01-01

    Listeria monocytogenes is the causative bacteria of listeriosis, which has a higher mortality rate than that of other causes of food poisoning. Listeria spp., of which L. monocytogenes is a member, have been isolated from food and manufacturing environments. Several methods have been published for identifying Listeria spp.; however, many of the methods cannot identify newly categorized Listeria spp. Additionally, they are often not suitable for the food industry, owing to their complexity, cost, or time consumption. Recently, high-resolution melting analysis (HRMA), which exploits DNA-sequence differences, has received attention as a simple and quick genomic typing method. In the present study, a new method for the simple, rapid, and low-cost identification of Listeria spp. has been presented using the genes rarA and ldh as targets for HRMA. DNA sequences of 9 Listeria species were first compared, and polymorphisms were identified for each species for primer design. Species specificity of each HRM curve pattern was estimated using type strains of all the species. Among the 9 species, 7 were identified by HRMA using rarA gene, including 3 new species. The remaining 2 species were identified by HRMA of ldh gene. The newly developed HRMA method was then used to assess Listeria isolates from the food industry, and the method efficiency was compared to that of identification by 16S rDNA sequence analysis. The 2 methods were in coherence for 92.6% of the samples, demonstrating the high accuracy of HRMA. The time required for identifying Listeria spp. was substantially low, and the process was considerably simplified, providing a useful and precise method for processing multiple samples per day. Our newly developed method for identifying Listeria spp. is highly valuable; its use is not limited to the food industry, and it can be used for the isolates from the natural environment.

  17. Tool for Rapid & Easy Identification of High Risk Diabetic Foot: Validation & Clinical Pilot of the Simplified 60 Second Diabetic Foot Screening Tool.

    Directory of Open Access Journals (Sweden)

    M Gail Woodbury

    Full Text Available Most diabetic foot amputations are caused by ulcers on the skin of the foot i.e. diabetic foot ulcers. Early identification of patients at high risk for diabetic foot ulcers is crucial. The 'Simplified 60-Second Diabetic Foot Screening Tool' has been designed to rapidly detect high risk diabetic feet, allowing for timely identification and referral of patients needing treatment. This study aimed to determine the clinical performance and inter-rater reliability of 'Simplified 60 Second Diabetic Foot Screening Tool' in order to evaluate its applicability for routine screening.The tool was independently tested by n=12 assessors with n=18 Guyanese patients with diabetes. Inter-rater reliability was assessed by calculating Cronbach's alpha for each of the assessment items. A minimum value of 0.60 was considered acceptable. Reliability scores of the screening tool assessment items were: 'monofilament test' 0.98; 'active ulcer' 0.97; 'previous amputation' 0.97; 'previous ulcer' 0.97; 'fixed ankle' 0.91; 'deformity' 0.87; 'callus' 0.87; 'absent pulses' 0.87; 'fixed toe' 0.80; 'blisters' 0.77; 'ingrown nail' 0.72; and 'fissures' 0.55. The item 'stiffness in the toe or ankle' was removed as it was observed in only 1.3% of patients. The item 'fissures' was also removed due to low inter-rater reliability. Clinical performance was assessed via a pilot study utilizing the screening tool on n=1,266 patients in an acute care setting in Georgetown, Guyana. In total, 48% of patients either had existing diabetic foot ulcers or were found to be at high risk for developing ulcers.Clinicians in low and middle income countries such as Guyana can use the Simplified 60-Second Diabetic Screening Tool to facilitate early detection and appropriate treatment of diabetic foot ulcers. Implementation of this screening tool has the potential to decrease diabetes related disability and mortality.

  18. Identification of potentially emerging food safety issues by analysis of reports published by the European Community's Rapid Alert System for Food and Feed (RASFF) during a four-year period

    NARCIS (Netherlands)

    Kleter, G.A.; Prandini, A.; Filippi, L.; Marvin, H.J.P.

    2009-01-01

    The SAFE FOODS project undertakes to design a new approach towards the early identification of emerging food safety hazards. This study explored the utility of notifications filed through RASFF, the European Commission¿s Rapid Alert System for Food and Feed, to identify emerging trends in food

  19. Evaluation of a PCR assay for identification and differentiation of Campylobacter fetus subspecies

    DEFF Research Database (Denmark)

    Hum, S.; Quinn, K.; Brunner, J.

    1997-01-01

    Objective To evaluate a polymerase chain reaction assay for identification of Campylobacter fetus and differentiation of the defined subspecies. Design Characterisation of bacterial strains by traditional phenotyping, polymerase chain reaction, a probabilistic identification scheme and macrorestr......Objective To evaluate a polymerase chain reaction assay for identification of Campylobacter fetus and differentiation of the defined subspecies. Design Characterisation of bacterial strains by traditional phenotyping, polymerase chain reaction, a probabilistic identification scheme...... by traditional phenotypic methods and the PCR assay was found to be 80.8%. The polymerase chain reaction proved to be a reliable technique for the species and subspecies identification of C fetus; equivocal results were obtained in only two instances. Initial misidentifications by conventional phenotyping...... methods were attributed to methodological differences used in various laboratories. Conclusion Our results indicate that misidentification of C fetus in routine diagnostic laboratories may be relatively common. The PCR assay evaluated gave rapid and reproducible results and is thus a valuable adjunctive...

  20. Performance of a PCR assay for the rapid identification of the Klebsiella pneumoniae ST258 epidemic clone in Latin American clinical isolates.

    Science.gov (United States)

    Gomez, S A; Rapoport, M; Piergrossi, N; Faccone, D; Pasteran, F; De Belder, D; ReLAVRA-Group; Petroni, A; Corso, A

    2016-10-01

    The worldwide dissemination of Klebsiella pneumoniae carbapenemase (KPC)-producing Klebsiella pneumoniae ST258 demands a rapid PCR-based typing method to detect unique genes of the ST258 clone. This study evaluates a PCR developed by Adler et al. (2014) for the detection of ST258 in K. pneumoniae clinical isolates centered on the identification of the pilv-I and prp genes. We tested 143 clinical isolates from Argentina (n=109), Chile (n=1), Colombia (n=1), Costa Rica (n=2), Ecuador (n=5), El Salvador (n=2), Nicaragua (n=5), Panamá (n=2), Paraguay (n=2), Perú (n=3) and Trinidad and Tobago (n=11) recovered from 2006 to 2015. blaKPC, pilv-l and prp genes were detected by PCR and sequenced by standard procedures. ST258 and non-ST258 were defined by PFGE and/or MLST. Isolates were grouped according to PFGE patterns: 58 were compatible with ST258 (group 1) and 85 with non-ST258 (group 2). MLST study was done on an arbitrary selection of isolates. The pilv-l gene was present only in ST258 isolates, regardless of the presence of the blaKPC gene. Results for the prp gene were variable. Its presence did not define ST258. The pilv-I PCR had a sensitivity and specificity of 100%, respectively, for the detection of ST258 in the isolates under investigation. Given our findings, the pilv-I PCR could replace more time and resource consuming methods, allowing for more rapid detection of the circulating high risk K. pneumoniae clone ST258 in Latin American (LA) countries. Copyright © 2016 Elsevier B.V. All rights reserved.

  1. Perry's Intellectual Scheme.

    Science.gov (United States)

    Love, Patrick G.; Guthrie, Victoria L.

    1999-01-01

    Summarizes William Perry's intellectual scheme and places it in the context of the 1990's. Perry's scheme of cognitive development, though more than thirty years old, is still being used by practitioners today to enhance practice in and out of the classroom. It laid a foundation for new research to extend, challenge, and build onto the scheme.…

  2. Finite Boltzmann schemes

    NARCIS (Netherlands)

    Sman, van der R.G.M.

    2006-01-01

    In the special case of relaxation parameter = 1 lattice Boltzmann schemes for (convection) diffusion and fluid flow are equivalent to finite difference/volume (FD) schemes, and are thus coined finite Boltzmann (FB) schemes. We show that the equivalence is inherent to the homology of the

  3. Rapid evaporative ionisation mass spectrometry of electrosurgical vapours for the identification of breast pathology: towards an intelligent knife for breast cancer surgery.

    Science.gov (United States)

    St John, Edward R; Balog, Julia; McKenzie, James S; Rossi, Merja; Covington, April; Muirhead, Laura; Bodai, Zsolt; Rosini, Francesca; Speller, Abigail V M; Shousha, Sami; Ramakrishnan, Rathi; Darzi, Ara; Takats, Zoltan; Leff, Daniel R

    2017-05-23

    Re-operation for positive resection margins following breast-conserving surgery occurs frequently (average = 20-25%), is cost-inefficient, and leads to physical and psychological morbidity. Current margin assessment techniques are slow and labour intensive. Rapid evaporative ionisation mass spectrometry (REIMS) rapidly identifies dissected tissues by determination of tissue structural lipid profiles through on-line chemical analysis of electrosurgical aerosol toward real-time margin assessment. Electrosurgical aerosol produced from ex-vivo and in-vivo breast samples was aspirated into a mass spectrometer (MS) using a monopolar hand-piece. Tissue identification results obtained by multivariate statistical analysis of MS data were validated by histopathology. Ex-vivo classification models were constructed from a mass spectral database of normal and tumour breast samples. Univariate and tandem MS analysis of significant peaks was conducted to identify biochemical differences between normal and cancerous tissues. An ex-vivo classification model was used in combination with bespoke recognition software, as an intelligent knife (iKnife), to predict the diagnosis for an ex-vivo validation set. Intraoperative REIMS data were acquired during breast surgery and time-synchronized to operative videos. A classification model using histologically validated spectral data acquired from 932 sampling points in normal tissue and 226 in tumour tissue provided 93.4% sensitivity and 94.9% specificity. Tandem MS identified 63 phospholipids and 6 triglyceride species responsible for 24 spectral differences between tissue types. iKnife recognition accuracy with 260 newly acquired fresh and frozen breast tissue specimens (normal n = 161, tumour n = 99) provided sensitivity of 90.9% and specificity of 98.8%. The ex-vivo and intra-operative method produced visually comparable high intensity spectra. iKnife interpretation of intra-operative electrosurgical vapours, including data

  4. Rapid and accurate species and genomic species identification and exhaustive population diversity assessment of Agrobacterium spp. using recA-based PCR.

    Science.gov (United States)

    Shams, M; Vial, L; Chapulliot, D; Nesme, X; Lavire, C

    2013-07-01

    Agrobacteria are common soil bacteria that interact with plants as commensals, plant growth promoting rhizobacteria or alternatively as pathogens. Indigenous agrobacterial populations are composites, generally with several species and/or genomic species and several strains per species. We thus developed a recA-based PCR approach to accurately identify and specifically detect agrobacteria at various taxonomic levels. Specific primers were designed for all species and/or genomic species of Agrobacterium presently known, including 11 genomic species of the Agrobacterium tumefaciens complex (G1-G9, G13 and G14, among which only G2, G4, G8 and G14 still received a Latin epithet: pusense, radiobacter, fabrum and nepotum, respectively), A. larrymoorei, A. rubi, R. skierniewicense, A. sp. 1650, and A. vitis, and for the close relative Allorhizobium undicola. Specific primers were also designed for superior taxa, Agrobacterium spp. and Rhizobiaceace. Primer specificities were assessed with target and non-target pure culture DNAs as well as with DNAs extracted from composite agrobacterial communities. In addition, we showed that the amplicon cloning-sequencing approach used with Agrobacterium-specific or Rhizobiaceae-specific primers is a way to assess the agrobacterial diversity of an indigenous agrobacterial population. Hence, the agrobacterium-specific primers designed in the present study enabled the first accurate and rapid identification of all species and/or genomic species of Agrobacterium, as well as their direct detection in environmental samples. Copyright © 2013 Elsevier GmbH. All rights reserved.

  5. Reduced Dimensionality tailored HN(C)N Pulse Sequences for Efficient Backbone Resonance Assignment of Proteins through Rapid Identification of Sequential HSQC peaks

    CERN Document Server

    Kumar, Dinesh

    2013-01-01

    Two novel reduced dimensionality (RD) experiments -(4,3)D-hNCOcaNH and (4,3)D-hNcoCANH- have been presented here to facilitate the backbone resonance assignment of proteins both in terms of its accuracy and speed. The experiments basically represent an improvisation of previously reported HN(C)N experiment [Panchal et. al., J. Biomol. NMR. (2002), 20 (2), 135-147] and exploit the simple reduced dimensionality NMR concept [Szyperski et. al. (2002), Proc. Natl. Acad. Sci. U.S.A. 99(12), 8009-8014] to achieve (a) higher dispersion and resolution along the co-evolved F1 dimension and (b) rapid identification of sequential HSQC peaks on its F2(15N)- F3(1H) planes. The current implementation is based on the fact that the linear combination of 15N and 13CO/13Ca chemical shifts offers relatively better dispersion and randomness compared to the individual chemical shifts; thus enables the assignment of crowded HSQC spectra by resolving the ambiguities generally encountered in HNCN based assignment protocol because of ...

  6. Multicenter Evaluation of the Portrait Staph ID/R Blood Culture Panel for Rapid Identification of Staphylococci and Detection of themecAGene.

    Science.gov (United States)

    Denys, Gerald A; Collazo-Velez, Vanessa; Young, Stephen; Daly, Judy A; Couturier, Marc Roger; Faron, Matthew L; Buchan, Blake W; Ledeboer, Nathan

    2017-04-01

    Bloodstream infections are a leading cause of morbidity and mortality in the United States and are associated with increased health care costs. We evaluated the Portrait Staph ID/R blood culture panel (BCP) multiplex PCR assay (Great Basin Scientific, Salt Lake City, UT) for the rapid and simultaneous identification (ID) of Staphylococcus aureus , Staphylococcus lugdunensis , and Staphylococcus species to the genus level and the detection of the mecA gene directly from a positive blood culture bottle. A total of 765 Bactec bottles demonstrating Gram-positive cocci in singles or clusters were tested during the prospective trial at 3 clinical sites. The Portrait Staph ID/R BCP results were compared with results from conventional biochemical and cefoxitin disk methods performed at an independent laboratory. Discordant ID and mecA results were resolved by rpoB gene sequencing and mecA gene sequencing, respectively. A total of 658 Staphylococcus species isolates ( S. aureus , 211 isolates; S. lugdunensis , 3 isolates; and Staphylococcus spp., 444 isolates) were recovered from monomicrobial and 33 polymicrobial blood cultures. After discrepant analysis, the overall ratios of Portrait Staph ID/R BCP positive percent agreement and negative percent agreement were 99.4%/99.9% for Staphylococcus ID and 99.7%/99.2% for mecA detection. Copyright © 2017 American Society for Microbiology.

  7. Fusarium keratitis in South India: causative agents, their antifungal susceptibilities and a rapid identification method for the Fusarium solani species complex.

    Science.gov (United States)

    Homa, Mónika; Shobana, Coimbatore S; Singh, Yendrembam R B; Manikandan, Palanisamy; Selvam, Kanesan P; Kredics, László; Narendran, Venkatapathy; Vágvölgyi, Csaba; Galgóczy, László

    2013-09-01

    Seventy Fusarium isolates derived from human keratomycosis were identified based on partial sequences of the β-tubulin (β-TUB) and translation elongation factor 1α (EF-1α) genes. Most of the isolates were confirmed as members of the F. solani species complex (75.71%), followed by the F. dimerum species complex (8.57%), the F. fujikuroi species complex (8.57%), the F. oxysporum species complex (4.29%) and the F. incarnatum-equiseti species complex (2.86%). A combined phylogenetic tree was estimated including all the 70 isolates. Isolates belonging to different species complexes formed separate clades. In this study, we also report the first isolation of F. napiforme from human keratomycosis. A new method based on a specific EcoRI restriction site in the EF-1α gene was developed for the rapid identification of F. solani. In vitro antifungal susceptibilities of the isolates to seven antifungals were determined by broth microdilution method. Terbinafine, natamycin and amphotericin B proved to be the most effective drugs, followed by voriconazole. The minimal inhibitory concentrations of clotrimazole, econazole and itraconazole were generally high (≥64 μg ml(-1) ). The interactions between the two most effective antifungals (natamycin and terbinafine) were determined by checkerboard microdilution method. Synergism (71.8%) or no interaction (28.2%) was revealed between the two compounds. © 2013 Blackwell Verlag GmbH.

  8. An integrated strategy for rapid discovery and identification of the sequential piperine metabolites in rats using ultra high-performance liquid chromatography/high resolution mass spectrometery.

    Science.gov (United States)

    Shang, Zhanpeng; Cai, Wei; Cao, Yanfeng; Wang, Fei; Wang, Zhibin; Lu, Jianqiu; Zhang, Jiayu

    2017-11-30

    Piperine, one of the major bioactive constituents isolated from natural flavorings and medicinal-culinary herbs, possesses various biological activities. In the present study, an integrated strategy based on ultra high-performance liquid chromatography/high resolution mass spectrometry was established to reveal piperine metabolism in rats. First of all, post-acquisition data-mining methods, including high resolution extracted ion chromatograms (HREICs) and multiple mass defect filtering (MMDF), were used to screen piperine metabolite candidates in a full-scan HRMS(1) level. Then, parent ion list-dynamic exclusion coupled with data-dependent data-acquisition method was utilized to acquire MS(n) datasets. In addition, the established reverse molecular assembly (RMA) approach based on paired diagnostic product ions (pDPIs) coupled with neutral loss fragments (NLFs) was used to ascertain and identify the major-to-trace piperine metabolites efficiently. And then, the calculated ClogP values were utilized to distinguish the positional isomers. As a result, a total of 148 piperine metabolites were detected and characterized tentatively. The results demonstrated that piperine mainly underwent hydrogenation, dehydrogenation, hydroxylation, glucuronide conjugation, sulfate conjugation, ring-cleavage, and their composite reactions. Our results not only provided novel and useful data to better understand the safety, toxicity and efficacy of this potential therapeutic agent, but also indicated that the proposed strategy was reliable for a rapid discovery and identification drug-related constituents in vivo. Copyright © 2017. Published by Elsevier B.V.

  9. Rapid Identification of Flavonoid Constituents Directly from PTP1B Inhibitive Extract of Raspberry (Rubus idaeus L.) Leaves by HPLC-ESI-QTOF-MS-MS.

    Science.gov (United States)

    Li, Zhuan-Hong; Guo, Han; Xu, Wen-Bin; Ge, Juan; Li, Xin; Alimu, Mireguli; He, Da-Jun

    2016-01-01

    Many potential health benefits of raspberry (Rubus idaeus L.) leaves were attributed to polyphenolic compounds, especially flavonoids. In this study, the methanol extract of R. idaeus leaves showed significant protein tyrosine phosphatase-1B (PTP1B) inhibitory activity with IC50 value of 3.41 ± 0.01 µg mL(-1) Meanwhile, a rapid and reliable method, employed high-performance liquid chromatography coupled with electrospray ionization quadrupole time-of-flight tandem mass spectrometry, was established for structure identification of flavonoids from PTP1B inhibitive extract of R. idaeus leaves using accurate mass measurement and characteristic fragmentation patterns. A total of 16 flavonoids, including 4 quercetin derivatives, 2 luteolin derivatives, 8 kaempferol derivatives and 2 isorhamnetin derivatives, were identified. Compounds 3: and 4: , Compounds 6: and 7: and Compounds 15: and 16: were isomers with different aglycones and different saccharides. Compounds 8: , 9: and 10: were isomers with the same aglycone and the same saccharide but different substituent positions. Compounds 11: and 12: were isomers with the same aglycone but different saccharides. Compounds 2: , 8: , 9: and 10: possessed the same substituent saccharide of glycuronic acid. Most of them were reported inR. idaeus for the first time. © The Author 2016. Published by Oxford University Press. All rights reserved. For Permissions, please email: journals.permissions@oup.com.

  10. Rapid Identification of Flavonoid Constituents Directly from PTP1B Inhibitive Extract of Raspberry (Rubus idaeus L.) Leaves by HPLC–ESI–QTOF–MS-MS

    Science.gov (United States)

    Li, Zhuan-Hong; Guo, Han; Xu, Wen-Bin; Ge, Juan; Li, Xin; Alimu, Mireguli; He, Da-Jun

    2016-01-01

    Many potential health benefits of raspberry (Rubus idaeus L.) leaves were attributed to polyphenolic compounds, especially flavonoids. In this study, the methanol extract of R. idaeus leaves showed significant protein tyrosine phosphatase-1B (PTP1B) inhibitory activity with IC50 value of 3.41 ± 0.01 µg mL−1. Meanwhile, a rapid and reliable method, employed high-performance liquid chromatography coupled with electrospray ionization quadrupole time-of-flight tandem mass spectrometry, was established for structure identification of flavonoids from PTP1B inhibitive extract of R. idaeus leaves using accurate mass measurement and characteristic fragmentation patterns. A total of 16 flavonoids, including 4 quercetin derivatives, 2 luteolin derivatives, 8 kaempferol derivatives and 2 isorhamnetin derivatives, were identified. Compounds 3 and 4, Compounds 6 and 7 and Compounds 15 and 16 were isomers with different aglycones and different saccharides. Compounds 8, 9 and 10 were isomers with the same aglycone and the same saccharide but different substituent positions. Compounds 11 and 12 were isomers with the same aglycone but different saccharides. Compounds 2, 8, 9 and 10 possessed the same substituent saccharide of glycuronic acid. Most of them were reported in R. idaeus for the first time. PMID:26896347

  11. A Rapid and Reproducible Genomic DNA Extraction Protocol for Sequence-Based Identification of Archaea, Bacteria, Cyanobacteria, Diatoms, Fungi, and Green Algae

    Directory of Open Access Journals (Sweden)

    Farkhondeh Saba

    2017-01-01

    Full Text Available Background:  Sequence-based identification of various microorganisms including Archaea, Bacteria, Cyanobacteria, Diatoms, Fungi, and green algae necessitates an efficient and reproducible genome extraction procedure though which a pure template DNA is yielded and it can be used in polymerase chain reactions (PCR. Considering the fact that DNA extraction from these microorganisms is time consuming and laborious, we developed and standardized a safe, rapid and inexpensive miniprep protocol. Methods:  According to our results, amplification of various genomic regions including SSU, LSU, ITS, β-tubulin, actin, RPB2, and EF-1 resulted in a reproducible and efficient DNA extraction from a wide range of microorganisms yielding adequate pure genomic material for reproducible PCR-amplifications. Results:   This method relies on a temporary shock of increased concentrations of detergent which can be applied concomitant with multiple freeze-thaws to yield sufficient amount of DNA for PCR amplification of multiple or single fragments(s of the genome. As an advantage, the recipe seems very flexible, thus, various optional steps can be included depending on the samples used.Conclusion:   Having the needed flexibility in each step, this protocol is applicable on a very wide range of samples. Hence, various steps can be included depending on the desired quantity and quality.

  12. Diagnostic accuracy of a standardized scheme for identification of Streptococcus uberis in quarter milk samples: A comparison between conventional bacteriological examination, modified Rambach agar medium culturing, and 16S rRNA gene sequencing.

    Science.gov (United States)

    Wald, Regina; Baumgartner, Martina; Urbantke, Verena; Stessl, Beatrix; Wittek, Thomas

    2017-02-01

    Bacteriological examination of milk samples is a prerequisite for pathogen-specific therapy and aids in limiting antimicrobial resistance. The aims of this study were to establish a standardized scheme for reliable Streptococcus uberis identification in routine diagnosis and to evaluate the accuracy of conventional tests and growing patterns of Strep. uberis on a selective medium (modified Rambach agar medium, MRAM) using 16S rRNA gene sequencing analysis as a reference method. We obtained isolates of presumptive Strep. uberis (n = 336) from quarter milk samples of dairy cows with intramammary infections and classified the isolates into 2 clusters using biochemical characterization. In cluster 1 (n = 280), cocci grew as non-hemolytic colonies, hydrolyzing esculin, carrying no Lancefield antigen (A/B/C/D/G) or Christie Atkins Munch-Petersen factor, and their growth was inhibited on an Enterococcus agar. Production of β-d-galactosidase on MRAM was shown by 257 of the cluster 1 isolates (91.79%), and 16S rRNA gene sequencing verified 271 (96.79%) of the isolates to be Strep. uberis. In 264 isolates (94.29%), MRAM agreed with the sequencing results. In cluster 2 (n = 56), isolates showed different characteristics: 37 (66.07%) were β-d-galactosidase-positive, and based on 16S sequencing results, 36 (64.29%) were identified correctly as Strep. uberis using biochemical methods. Identification success in this group differed significantly between routine diagnosis and MRAM application: MRAM agreed with sequencing results in 47 isolates (83.93%). To identify Strep. uberis and differentiate it from other lactic acid bacteria in routine diagnosis, we suggest using catalase reaction, hemolysis, esculin hydrolysis, and growth on enterococci agar. Isolates that show a typical biochemical profile can be identified satisfactorily with these tests. For Strep. uberis isolates with divergent patterns, application of MRAM as a follow-up test increased the diagnostic accuracy to 94

  13. Phenotypic identification of Porphyromonas gingivalis validated with matrix-assisted laser desorption/ionization time-of-flight mass spectrometry

    NARCIS (Netherlands)

    Rams, Thomas E; Sautter, Jacqueline D; Getreu, Adam; van Winkelhoff, Arie J

    OBJECTIVE: Porphyromonas gingivalis is a major bacterial pathogen in human periodontitis. This study used matrix-assisted laser desorption/ionization time-of-flight (MALDI-TOF) mass spectrometry to assess the accuracy of a rapid phenotypic identification scheme for detection of cultivable P.

  14. Rapid identification and quantification of methamphetamine and amphetamine in hair by gas chromatography/mass spectrometry coupled with micropulverized extraction, aqueous acetylation and microextraction by packed sorbent.

    Science.gov (United States)

    Miyaguchi, Hajime; Iwata, Yuko T; Kanamori, Tatsuyuki; Tsujikawa, Kenji; Kuwayama, Kenji; Inoue, Hiroyuki

    2009-05-01

    We developed a rapid identification and quantification method for the toxicological analysis of methamphetamine and amphetamine in human hair by gas chromatography/mass spectrometry coupled with a novel combination of micropulverized extraction, aqueous acetylation and microextraction by packed sorbent (MEPS) named MiAMi-GC/MS. A washed hair sample (1-5 mg) was micropulverized for 5 min in a 2 mL plastic tube with 250 microL of water. An anion-exchange sorbent was added to adsorb anionic interferences. After removing the residue with a membrane-filter unit, sodium carbonate and acetic anhydride was admixed in turn. Acetylation was completed in approximately 20 min at room temperature. The acetylated analytes in the reaction liquid were concentrated to an octadecylsilica sorbent packed in the needle of a syringe by a CombiPAL autosampler. Elution was carried out with 50 microL of methanol, and the entire eluate injected into a gas chromatograph using a programmable temperature vaporizing (PTV) technique. The time required for sample preparation and GC/MS analysis was approximately 1 h from a washed hair sample, and an evaporation process was not required. Ranges for quantification were 0.20-50 (ng/mg) each for methamphetamine and amphetamine using 1 mg of hair. Accuracy and relative standard deviation (RSD) were evaluated intraday and interday at three concentrations, and the results were within the limit of a guidance issued by U.S. Food and Drug Administration. For identification, full-scan mass spectra of methamphetamine and amphetamine were obtained using 5 mg of fortified hair samples at 0.2 ng/mg. The extraction device of MEPS was durable for at least 300 extractions, whereas the liner of the gas chromatograph should be replaced after 20-30 times use. The carry over was estimated to be about 1-2%. This sample-preparation method coupled with GC/MS is fast and labor-saving in comparison with conventional methods.

  15. Quality of Care Is Improved by Rapid Short Incubation MALDI-ToF Identification from Blood Cultures as Measured by Reduced Length of Stay and Patient Outcomes as Part of a Multi-Disciplinary Approach to Bacteremia in Pediatric Patients.

    Directory of Open Access Journals (Sweden)

    Johannes A Delport

    Full Text Available Sepsis has seen an incremental increase in cases of about 13% annually in the USA and accounts for approximately 4400 deaths among pediatric patients. Early identification of the specific pathogen allows the clinician to ensure that the antibiotic coverage is optimal, an intervention that has been shown to improve patient outcomes in sepsis. Our study's objective was to assess the impact of a rapid Bruker MALDI-Tof identification protocol on pediatric sepsis cases by assessing various indicators. We assessed the quality of care by measuring the following indicators; time to identification of the pathogen, initiation of the most appropriate antibiotic, length of stay (LOS in hospital and patient outcomes, using a retrospective review over three consecutive years. In total 92 pediatric patients, similar in age and gender distributions were assessed; 37 in 2012, 33 in 2013 and 22 in 2014. The introduction of MALDI-TOF identification in 2013 led to a significant decrease in time to identify a pathogen by 21.03 hours (p = 1.95E-05. A short incubation MALDI-TOF identification protocol in 2014 further reduced time to identification by 17.75 hours (p = 2.48E-3. Overall in 2014 this led to a trend to earlier optimization of antibiotics by 20.2 hours (p = 0.14 and a reduction in length of stay after the implementation of MALDI-ToF identification in 2013 of 3.07 days and a further reduction of 8.92 days after the introduction of the rapid short incubation identification protocol using MALDI-Tof in 2014 (P = 0.12. By evaluating the subgroup of patients where antibiotics were changed, our study confirmed that patients received appropriate therapy 48.8% (20.2 hours earlier compared to conventional methods leading to a decrease in length of stay of 23.65 days after the implementation of MALDI-ToF identification and a further reduction of 9.82 days in 2014 compared to 2012 (p = 0.02. In 2014 outcomes between the patients needing a change in their antibiotic

  16. The Mawala irrigation scheme

    OpenAIRE

    de Bont, Chris

    2018-01-01

    This booklet was written to share research results with farmers and practitioners in Tanzania. It gives a summary of the empirical material collected during three months of field work in the Mawala irrigation scheme (Kilimanjaro Region), and includes maps, tables and photos. It describes the history of the irrigation scheme, as well current irrigation and farming practices. It especially focuses on the different kinds of infrastructural improvement in the scheme (by farmers and the government...

  17. Polymorphism-specific PCR enhances the diagnostic performance of American tegumentary leishmaniasis and allows the rapid identification of Leishmania species from Argentina

    Directory of Open Access Journals (Sweden)

    Marco Jorge D

    2012-08-01

    Full Text Available Abstract Background The diagnosis of the leishmaniases poses enormous challenges in Argentina. The Polymorphism-Specific PCR (PS-PCR designed and validated in our laboratories has been proven effective for typifying the Leishmania genus from cultured material. Here we evaluated the performance of this method in the diagnosis of American tegumentary leishmaniasis (ATL and the rapid identification of Leishmania spp. directly from clinical specimens. Methods A total of 63 patients from northwestern Argentina, with cutaneous or mucocutaneous lesions, underwent an ATL diagnosis protocol which included clinical examination, Leishmanin skin test, and microscopic examination of dermal smears. In addition, we performed PS-PCR on DNA directly extracted from the specimens scraped from the lesions. Results Out of the 63 patients, 44 were classified as ATL cases and 19 as non-ATL cases. The diagnostic sensitivity of the microscopic analysis of dermal smears and PS-PCR individually were 70.5% and 81%, respectively. When performing both tests in parallel, this parameter increased significantly to 97.6% (p = 0.0018. The specificities, on the other hand, were 100%, 84.2%, and 83.3% for the combination, respectively (p > 0.05. Using the PS-PCR analysis we successfully identified the Leishmania spp. in 31 out of the 44 ATL cases. Twenty-eight (90.3% cases were caused by L. (V. braziliensis, two (6.5% by L. (V. guyanensis, and one (3.2% by L. (V. panamensis. Conclusions The efficacy of the ATL diagnosis was significantly improved by combining the dermal smear examination with a PS-PCR analysis. Our strategy allowed us to reach the diagnosis of ATL with high accuracy regarding the species of the etiological agent in 70.5% of the cases. Moreover, we diagnosed two cases of the disseminated cutaneous form caused by L. (V. braziliensis and a cutaneous case due to L. (V. panamensis infection, both findings reported for the first time in Argentina.

  18. Polymorphism-specific PCR enhances the diagnostic performance of American tegumentary leishmaniasis and allows the rapid identification of Leishmania species from Argentina.

    Science.gov (United States)

    Marco, Jorge D; Barroso, Paola A; Mimori, Tatsuyuki; Locatelli, Fabricio M; Tomatani, Ayako; Mora, María C; Cajal, S Pamela; Nasser, Julio R; Parada, Luis A; Taniguchi, Taketoshi; Korenaga, Masataka; Basombrío, Miguel A; Hashiguchi, Yoshihisa

    2012-08-15

    The diagnosis of the leishmaniases poses enormous challenges in Argentina. The Polymorphism-Specific PCR (PS-PCR) designed and validated in our laboratories has been proven effective for typifying the Leishmania genus from cultured material. Here we evaluated the performance of this method in the diagnosis of American tegumentary leishmaniasis (ATL) and the rapid identification of Leishmania spp. directly from clinical specimens. A total of 63 patients from northwestern Argentina, with cutaneous or mucocutaneous lesions, underwent an ATL diagnosis protocol which included clinical examination, Leishmanin skin test, and microscopic examination of dermal smears. In addition, we performed PS-PCR on DNA directly extracted from the specimens scraped from the lesions. Out of the 63 patients, 44 were classified as ATL cases and 19 as non-ATL cases. The diagnostic sensitivity of the microscopic analysis of dermal smears and PS-PCR individually were 70.5% and 81%, respectively. When performing both tests in parallel, this parameter increased significantly to 97.6% (p = 0.0018). The specificities, on the other hand, were 100%, 84.2%, and 83.3% for the combination, respectively (p > 0.05). Using the PS-PCR analysis we successfully identified the Leishmania spp. in 31 out of the 44 ATL cases. Twenty-eight (90.3%) cases were caused by L. (V.) braziliensis, two (6.5%) by L. (V.) guyanensis, and one (3.2%) by L. (V.) panamensis. The efficacy of the ATL diagnosis was significantly improved by combining the dermal smear examination with a PS-PCR analysis. Our strategy allowed us to reach the diagnosis of ATL with high accuracy regarding the species of the etiological agent in 70.5% of the cases. Moreover, we diagnosed two cases of the disseminated cutaneous form caused by L. (V.) braziliensis and a cutaneous case due to L. (V.) panamensis infection, both findings reported for the first time in Argentina.

  19. Application of host-specific source-tracking tools for rapid identification of fecal contamination in fresh produce by humans and livestock.

    Science.gov (United States)

    Lee, Chang Soo; Lee, Jiyoung

    2013-03-30

    Fecal contamination in fresh produce is a public health concern because it may contain human pathogens. We introduced host-specific quantitative real-time polymerase chain reaction (qPCR) assays for the rapid detection and identification of fecal contamination sources from humans and farm animals (cow, pig, chicken) in fresh produce. Each composite fecal sample was spiked on lettuce at two contamination levels (0.2 mg or 2 mg feces g⁻¹), followed by qPCR assays for detecting each host-specific genetic marker: BoBac (cow); PF163 (pig); CP3-49 (chicken); and HF183 and gyrB (human). Two commercial DNA extraction kits were compared to evaluate DNA recovery yields and removal of PCR inhibition. Sketa2 assay was conducted to assess the presence of PCR inhibition in the contaminated lettuce. All the qPCR assays yielded reliable detection from contaminated lettuce (2 mg feces g⁻¹), where their target gene numbers were 1.5-5.0 × 10³ (HF183), 0.8-2.2 × 10³ (gyrB), 0.6-1.6 × 10³ (BoBac), 1.6-3.0 × 10³ (CP3-49) and 1.1-2.2 × 10³ (PF163) copies g⁻¹ of lettuce. Among the two extraction kits, QIAamp DNA Stool Kit resulted in 2-3 times higher sensitivity and 20% less PCR inhibition than the PowerFood™ kit. This study provides information on the optimized host-specific qPCR assay in identifying sources of fecal contamination in fresh produce and is useful for tracking the contamination source and improving agricultural practice. © 2012 Society of Chemical Industry.

  20. Genotype-specific real-time PCR combined with high-resolution melting analysis for rapid identification of red-spotted grouper nervous necrosis virus.

    Science.gov (United States)

    Toubanaki, Dimitra K; Karagouni, Evdokia

    2017-08-01

    A real-time genotype-specific polymerase chain reaction (PCR) assay combined with high-resolution melting (HRM) analysis was developed to assess the most common genotypes of nervous necrosis viruses or nodaviruses. Nodaviruses are the causal agents of viral nervous necrosis infections, which have been wreaking havoc in the aquaculture industry worldwide, with fish mortality up to 100%. The four different genotypes of nodaviruses correlate with differences in viral pathogenicity. Therefore, rational development of effective vaccines and diagnostics requires analysis of genetic variation among viruses. The aim of the present study was to develop a real-time tetra-primer genotype-specific PCR assay for genotype identification. Four primers were utilized for simultaneous amplification of nodavirus genotype-specific products in a single closed-tube PCR after a reverse-transcription reaction using RNA isolated from fish samples. For high-throughput sample analysis, SYBR Green-based real-time PCR was used in combination with HRM analysis. The assay was evaluated in terms of specificity and sensitivity. The analysis resulted in melting curves that were indicative of each genotype. The detection limit when using reference plasmids was 100 ag/µL for both genotypes, while the sensitivity of the assays when testing a complex mixture was 10 fg/µL for red-spotted grouper nervous necrosis virus (RGNNV) and 100 fg/µL for striped jack nervous necrosis virus (SJNNV). To test the capability of this method under real-world conditions, 58 samples were examined. All samples belonged to the RGNNV genotype, which was fully validated. The results were in full agreement with genotyping by reference methods. The proposed methodology provides a rapid, sensitive, specific, robust and automatable assay for nodavirus genotyping, making it a useful tool for diagnosis and screening for epidemiological studies.

  1. Optimization and validation of a fast amplification protocol for AmpFlSTR® Profiler Plus® for rapid forensic human identification.

    Science.gov (United States)

    Laurin, Nancy; Frégeau, Chantal

    2012-01-01

    The goal of this work was to optimize and validate a fast amplification protocol for the multiplex amplification of the STR loci included in AmpFlSTR(®) Profiler Plus(®) to expedite human DNA identification. By modifying the cycling conditions and by combining the use of a DNA polymerase optimized for high speed PCR (SpeedSTAR™ HS) and a more efficient thermal cycler instrument (Bio-RAD C1000™), we were able to reduce the amplification process from 4h to 26 min. No modification to the commercial AmpFlSTR(®) Profiler Plus(®) primer mix was required. When compared to the current Royal Canadian Mounted Police (RCMP) amplification protocol, no differences with regards to specificity, sensitivity, heterozygote peak height ratios and overall profile balance were noted. Moreover, complete concordance was obtained with profiles previously generated with the standard amplification protocol and minor alleles in mixture samples were reliably typed. An increase in n-4 stutter ratios (2.2% on average for all loci) was observed for profiles amplified with the fast protocol compared to the current procedure. Our results document the robustness of this rapid amplification protocol for STR profiling using the AmpFlSTR(®) Profiler Plus(®) primer set and demonstrate that comparable data can be obtained in substantially less time. This new approach could provide an alternative option to current multiplex STR typing amplification protocols in order to increase throughput or expedite time-sensitive cases. Copyright © 2011 Elsevier Ireland Ltd. All rights reserved.

  2. Impact of rapid microbial identification directly from positive blood cultures using matrix-assisted laser desorption/ionization time-of-flight mass spectrometry on patient management.

    Science.gov (United States)

    Martiny, D; Debaugnies, F; Gateff, D; Gérard, M; Aoun, M; Martin, C; Konopnicki, D; Loizidou, A; Georgala, A; Hainaut, M; Chantrenne, M; Dediste, A; Vandenberg, O; Van Praet, S

    2013-12-01

    For septic patients, delaying the initiation of antimicrobial therapy or choosing an inappropriate antibiotic can considerably worsen their prognosis. This study evaluated the impact of rapid microbial identification (RMI) from positive blood cultures on the management of patients with suspected sepsis. During a 6-month period, RMI by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) was performed for all new episodes of bacteraemia. For each patient, the infectious disease specialist was contacted and questioned about his therapeutic decisions made based on the Gram staining and the RMI. This information was collected to evaluate the number of RMIs that led to a therapeutic change or to a modification of the patient's general management (e.g. fast removal of infected catheters). During the study period, 277 new episodes of bacteraemia were recorded. In 71.12% of the cases, MALDI-TOF MS resulted in a successful RMI (197/277). For adult and paediatric patients, 13.38% (21/157) and 2.50% (1/40) of the RMIs, respectively, resulted in modification of the treatment regimen, according to the survey. In many other cases, the MALDI-TOF MS was a helpful tool for infectious disease specialists because it confirmed suspected cases of contamination, especially in the paediatric population (15/40 RMIs, 37.50%), or suggested complementary diagnostic testing. This study emphasizes the benefits of RMI from positive blood cultures. Although the use of this technique represents an extra cost for the laboratory, RMI using MALDI-TOF MS has been implemented in our daily practice. © 2013 The Authors Clinical Microbiology and Infection © 2013 European Society of Clinical Microbiology and Infectious Diseases.

  3. Towards Symbolic Encryption Schemes

    DEFF Research Database (Denmark)

    Ahmed, Naveed; Jensen, Christian D.; Zenner, Erik

    2012-01-01

    , namely an authenticated encryption scheme that is secure under chosen ciphertext attack. Therefore, many reasonable encryption schemes, such as AES in the CBC or CFB mode, are not among the implementation options. In this paper, we report new attacks on CBC and CFB based implementations of the well...

  4. Multiresolution signal decomposition schemes

    NARCIS (Netherlands)

    J. Goutsias (John); H.J.A.M. Heijmans (Henk)

    1998-01-01

    textabstract[PNA-R9810] Interest in multiresolution techniques for signal processing and analysis is increasing steadily. An important instance of such a technique is the so-called pyramid decomposition scheme. This report proposes a general axiomatic pyramid decomposition scheme for signal analysis

  5. Adaptive protection scheme

    Directory of Open Access Journals (Sweden)

    R. Sitharthan

    2016-09-01

    Full Text Available This paper aims at modelling an electronically coupled distributed energy resource with an adaptive protection scheme. The electronically coupled distributed energy resource is a microgrid framework formed by coupling the renewable energy source electronically. Further, the proposed adaptive protection scheme provides a suitable protection to the microgrid for various fault conditions irrespective of the operating mode of the microgrid: namely, grid connected mode and islanded mode. The outstanding aspect of the developed adaptive protection scheme is that it monitors the microgrid and instantly updates relay fault current according to the variations that occur in the system. The proposed adaptive protection scheme also employs auto reclosures, through which the proposed adaptive protection scheme recovers faster from the fault and thereby increases the consistency of the microgrid. The effectiveness of the proposed adaptive protection is studied through the time domain simulations carried out in the PSCAD⧹EMTDC software environment.

  6. Apolipoprotein e4 Is Associated with More Rapid Decline in Odor Identification than in Odor Threshold or Dementia Rating Scale Scores

    Science.gov (United States)

    Calhoun-Haney, R.; Murphy, C.

    2005-01-01

    Individuals with the apolipoprotein E e4 genetic risk factor for Alzheimer's disease (AD) show deficits in olfactory function. The purpose of the present study was to examine longitudinally odor identification (odor ID), odor threshold, picture identification, and global cognitive status in allele positive (e4+) and negative (e4-) persons.…

  7. Rapid identification of microorganisms from positive blood cultures by testing early growth on solid media using matrix-assisted laser desorption ionization-time of flight mass spectrometry.

    Science.gov (United States)

    Gonzalez, Mark D; Weber, Carol J; Burnham, Carey-Ann D

    2016-06-01

    We performed a retrospective analysis of a simple modification to MALDI-TOF MS for microorganism identification to accurately improve the turnaround time (TAT) for identification of Enterobacteriaceae recovered in blood cultures. Relative to standard MALDI-TOF MS procedures, we reduced TAT from 28.3 (n=90) to 21.2h (n=107). Copyright © 2016 Elsevier Inc. All rights reserved.

  8. Parallel particle identification and separation for active optical sorting

    DEFF Research Database (Denmark)

    Perch-Nielsen, Ivan R.; Palima, Darwin; Dam, Jeppe Seidelin

    2009-01-01

    An instrument for rapidly and non-invasively sorting different cell specimens is a valuable tool in biological and medical research. Parallel identification of target specimens through image analysis can sort based on highly tuneable selection criteria and can enable high-speed optical sorting when...... matched with a rapidly reconfigurable optical sorting field. We demonstrate the potential of such a system using colloidal polystyrene microspheres. By combining machine vision with a parallel add-on optical manipulation scheme, we were able to move identified particles over a distance of several hundred...

  9. Succesful labelling schemes

    DEFF Research Database (Denmark)

    Juhl, Hans Jørn; Stacey, Julia

    2001-01-01

    It is usual practice to evaluate the success of a labelling scheme by looking at the awareness percentage, but in many cases this is not sufficient. The awareness percentage gives no indication of which of the consumer segments that are aware of and use labelling schemes and which do not....... In the spring of 2001 MAPP carried out an extensive consumer study with special emphasis on the Nordic environmentally friendly label 'the swan'. The purpose was to find out how much consumers actually know and use various labelling schemes. 869 households were contacted and asked to fill in a questionnaire....... 664 households returned a completed questionnaire. There were five answering categories for each label in the questionnaire: * have not seen the label before. * I have seen the label before but I do not know the precise contents of the labelling scheme. * I have seen the label before, I do not know...

  10. Tabled Execution in Scheme

    Energy Technology Data Exchange (ETDEWEB)

    Willcock, J J; Lumsdaine, A; Quinlan, D J

    2008-08-19

    Tabled execution is a generalization of memorization developed by the logic programming community. It not only saves results from tabled predicates, but also stores the set of currently active calls to them; tabled execution can thus provide meaningful semantics for programs that seemingly contain infinite recursions with the same arguments. In logic programming, tabled execution is used for many purposes, both for improving the efficiency of programs, and making tasks simpler and more direct to express than with normal logic programs. However, tabled execution is only infrequently applied in mainstream functional languages such as Scheme. We demonstrate an elegant implementation of tabled execution in Scheme, using a mix of continuation-passing style and mutable data. We also show the use of tabled execution in Scheme for a problem in formal language and automata theory, demonstrating that tabled execution can be a valuable tool for Scheme users.

  11. Iris feature based watermarking algorithm for personal identification

    Science.gov (United States)

    Feng, Gui; Lin, Qiwei

    2007-11-01

    Rapidly development of technique causes big problem on the security of digital product. The digital signature technique and watermarking technique are common used solution, but hidden security trouble is still existed. A new biometric watermarking scheme is proposed for property protecting and digital product owner identification. The scheme adopts the iris of digital product holder as watermark, which has the advantage of uniqueness, stability and impossible been counterfeited, so the security level is improved. In this scheme, we utilize person's iris to generate the watermark, and embed this watermark signal into the host image like certification of consumer or other document. While identifying or verifying the owner of the product, we first extract and reconstruct the watermark, then compare the watermark (i.e. the iris image) with the field collected iris image to verify authority. Because there is a direct relationship between the certification holder and the credentials; the reliability is improved greatly.

  12. Extra pulmonary tuberculosis: Rapid identification of Mycobacterium tuberculosis grown in Mycobacterium growth indicator tube 960 and Lowenstein-Jensen media, employing Standard diagnostics Bioline Mycobacterium tuberculosis protein 64 antigen detection kit

    Directory of Open Access Journals (Sweden)

    G Kandhakumari

    2015-01-01

    Full Text Available Background: Investigation of extra pulmonary tuberculosis (EPTB in and around Pondicherry is being carried out since August 2011 in our tertiary care super specialty hospital. Objectives: To compare the rapid Kit SD Bio-Line MPT 64 Ag with conventional and time consuming biochemical tests. Confirmation of Mycobacterium tuberculosis at a reasonable time frame is the main thrust. Materials and Methods: Thirty three Mycobacterium tuberculosis and four Non-Tuberculous Mycobacteria (NTM grown in MGIT960 system/Lowenstein-Jensen media (LJ were examined by the rapid MPT 64 antigen detection as well as a battery of conventional tests like niacin, nitrate reduction, paraminobenzoic acid susceptibility and cord formation. Results and Conclusion: . Both the rapid kit and conventional tests correctly identified 33 M.tuberculosis isolates. Keeping conventional identification as reference, sensitivity and specificity for rapid kit was 100%. Rapid kit which takes only 15 minutes is accurate, cost effective, and facilitates early treatment for these EPTB patients, whose clinical specimens are paucibacillary.

  13. Matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF) directly from positive blood culture flasks allows rapid identification of bloodstream infections in immunosuppressed hosts.

    Science.gov (United States)

    Egli, A; Osthoff, M; Goldenberger, D; Halter, J; Schaub, S; Steiger, J; Weisser, M; Frei, R

    2015-06-01

    In immunosuppressed hosts, rapid identification of microorganisms of bloodstream infections is crucial to ensuring effective antimicrobial therapy. Conventional culture requires up to 72 h from sample collection to pathogen identification. We used the SepsiTyper Kit and matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF; Microflex, Bruker) directly from positive blood culture (BacT/ALERT 3D, FN/FA vials; bioMérieux) in comparison to standard culture methodology (VITEK 2; bioMérieux) for species identification. A total of 62 consecutive positive blood cultures from immunosuppressed patients (solid organ or hematopoietic transplant recipients, or with febrile neutropenia) were analyzed. Culture yielded gram-negative bacteria (GNB) in 27/62 (43.5%) and gram-positive (GPB) in 35/62 (56.5%) vials. For GNB, the predominant species identified by MALDI-TOF and confirmed by VITEK were Escherichia coli (16/16 correctly identified) and Enterobacter cloacae (4/4), with a sensitivity and specificity of 92.6% and 100%, respectively. For GPB, predominant species were Staphylococcus aureus (3/3), coagulase-negative staphylococci (12/24), and Enterococcus faecium (6/6) with a sensitivity of 100%, 60%, and 100%, respectively. The median time from blood collection to species identification was 27.4 h with MALDI-TOF identification and 46.6 h with conventional methodology. Using MALDI-TOF directly from positive blood cultures allowed a shorter time to identification with high sensitivity and specificity in immunosuppressed patients. © 2015 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

  14. Compact Spreader Schemes

    Energy Technology Data Exchange (ETDEWEB)

    Placidi, M.; Jung, J. -Y.; Ratti, A.; Sun, C.

    2014-07-25

    This paper describes beam distribution schemes adopting a novel implementation based on low amplitude vertical deflections combined with horizontal ones generated by Lambertson-type septum magnets. This scheme offers substantial compactness in the longitudinal layouts of the beam lines and increased flexibility for beam delivery of multiple beam lines on a shot-to-shot basis. Fast kickers (FK) or transverse electric field RF Deflectors (RFD) provide the low amplitude deflections. Initially proposed at the Stanford Linear Accelerator Center (SLAC) as tools for beam diagnostics and more recently adopted for multiline beam pattern schemes, RFDs offer repetition capabilities and a likely better amplitude reproducibility when compared to FKs, which, in turn, offer more modest financial involvements both in construction and operation. Both solutions represent an ideal approach for the design of compact beam distribution systems resulting in space and cost savings while preserving flexibility and beam quality.

  15. Development of a Rapid and Accurate Identification Method for Citrobacter Species Isolated from Pork Products Using a Matrix-Assisted Laser-Desorption Ionization Time-of-Flight Mass Spectrometry (MALDI-TOF MS).

    Science.gov (United States)

    Kwak, Hye-Lim; Han, Sun-Kyung; Park, Sunghoon; Park, Si Hong; Shim, Jae-Yong; Oh, Mihwa; Ricke, Steven C; Kim, Hae-Yeong

    2015-09-01

    Previous detection methods for Citrobacter are considered time consuming and laborious. In this study, we have developed a rapid and accurate detection method for Citrobacter species in pork products, using matrix-assisted laser desorption ionization time-of-flight (MALDI-TOF) mass spectrometry (MS). A total of 35 Citrobacter strains were isolated from 30 pork products and identified by both MALDI-TOF MS and 16S rRNA gene sequencing approaches. All isolates were identified to the species level by the MALDI-TOF MS, while 16S rRNA gene sequencing results could not discriminate them clearly. These results confirmed that MALDI-TOF MS is a more accurate and rapid detection method for the identification of Citrobacter species.

  16. Neutral fragment filtering for rapid identification of new diester-diterpenoid alkaloids in roots of Aconitum carmichaeli by ultra-high-pressure liquid chromatography coupled with linear ion trap-orbitrap mass spectrometry.

    Directory of Open Access Journals (Sweden)

    Jing Zhang

    Full Text Available A rapid and effective method was developed for separation and identification of diester-diterpenoid alkaloids (DDA in the roots of Aconitum carmichaeli by ultra-high-pressure liquid chromatography coupled with high resolution LTQ-Orbitrap tandem mass spectrometry (UHPLC-LTQ-Orbitrap-MS(n. According to accurate mass measurement and the characteristic neutral loss filtering strategy, a total of 42 diester-diterpenoid alkaloids (DDA were rapidly detected and characterized or tentatively identified. Meanwhile, the proposed fragmentation pathways and the major diagnostic fragment ions of aconitine, mesaconitine and hypaconitine were investigated to trace DDA derivatives in crude plant extracts. 23 potential new compounds were successfully screened and characterized in Aconitum carmichaeli, including 16 short chain fatty acyls DDA, 4 N-dealkyl DDA and several isomers of aconitine, mesaconitine and hypaconitine.

  17. Validation of the 16S rDNA and COI DNA barcoding technique for rapid molecular identification of stored product psocids (Insecta: Psocodea: Liposcelididae).

    Science.gov (United States)

    Yang, Qianqian; Zhao, Shuo; Kucerová, Zuzana; Stejskal, Václav; Opit, George; Qin, Meng; Cao, Yang; Li, Fujun; Li, Zhihong

    2013-02-01

    Psocids are serious storage pests, and their control is hampered by the fact that different species respond differently to insecticides used for the control of stored-product insect pests. Additionally, psocids of genus Liposcelis that are commonly associated with stored-products are difficult to identify using morphological characteristics. The goal of this study was to validate molecular identification of stored-product psocids of genus Liposcelis based on 16S rDNA and cytochrome oxidase I (COI) DNA barcoding. Unidentified liposcelids (Liposcelis DK) imported from Denmark to China were compared with 14 population samples of seven common species (L. bostrychophila, L. brunnea, L. corrodens, L. decolor, L. entomophila, L. mendax, and L. paeta). The explored species (DK) liposcelids shared >98% sequence similarity for both the 16S rDNA and COI genes with the reference L. corrodens samples (98.32 and 98.94% for 16S rDNA and COI, respectively). A neighbor-joining tree revealed that the explored DK sample and the reference L. corrodens samples belong to the same clade. These molecular results were verified by morphological identification of DK specimens, facilitated by SEM microphotography. The DNA barcoding method and the neighbor-joining phylogenetic analyses indicated that both the 16S rDNA and COI genes were suitable for Liposcelis species identification. DNA barcoding has great potential for use in fast and accurate liposcelid identification.

  18. Phenotypic Identification of Actinomyces and Related Species Isolated from Human Sources

    Science.gov (United States)

    Sarkonen, Nanna; Könönen, Eija; Summanen, Paula; Könönen, Mauno; Jousimies-Somer, Hannele

    2001-01-01

    Recent advancements in chemotaxonomic and molecular biology-based identification methods have clarified the taxonomy of the genus Actinomyces and have led to the recognition of several new Actinomyces and related species. Actinomyces-like gram-positive rods have increasingly been isolated from various clinical specimens. Thus, an easily accessible scheme for reliable differentiation at the species level is needed in clinical and oral microbiology laboratories, where bacterial identification is mainly based on conventional biochemical methods. In the present study we designed a two-step protocol that consists of a flowchart that describes rapid, cost-efficient tests for preliminary identification of Actinomyces and closely related species and an updated more comprehensive scheme that also uses fermentation reactions for accurate differentiation of Actinomyces and closely related species. PMID:11682514

  19. Rapid identification of microorganisms by mass spectrometry: improved performance by incorporation of in-house spectral data into a commercial database.

    Science.gov (United States)

    Sogawa, Kazuyuki; Watanabe, Masaharu; Sato, Kenichi; Segawa, Syunsuke; Miyabe, Akiko; Murata, Syota; Saito, Tomoko; Nomura, Fumio

    2012-06-01

    Matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) is increasingly used as a microbial diagnostic method for species identification of pathogens. However, MALDI-TOF identification of bacteria at the species level remains unsatisfactory, with the major problem being an incomplete database that still needs refinement and expansion. Augmentation of the original MALDI BioTyper 2.0 (Bruker) database by incorporating mass spectra obtained in-house from clinical isolates may increase the identification rate at the species level. We conducted a prospective study to assess whether the augmented database can improve the performance of MALDI-TOF MS for routine identification of species. Cluster analyses revealed distinct differences in MS spectral profiles of clinical isolates obtained in our hospital and those of ATCC strains in the Bruker database. In the first part of the study, which was performed over 3 weeks, 259 bacterial isolates were subjected to analysis by MALDI-TOF MS, and MS spectra of 229 successfully identified isolates (49 species) were incorporated into the original database to give the augmented Bruker-Chiba database. In a second separate analysis, the concordance of identification of 498 clinical isolates of the 49 species with conventional methods was 87.1% (434/498) with the commercial Bruker database and 98.0% (488/498) using the Bruker-Chiba database. These results indicate that refinement of a commercial database can be achieved relatively easy and effectively by incorporating MS spectra of clinical isolates obtained in a clinical laboratory.

  20. Improving uptake and use of malaria rapid diagnostic tests in the context of artemisinin drug resistance containment in eastern Myanmar: an evaluation of incentive schemes among informal private healthcare providers.

    Science.gov (United States)

    Aung, Tin; White, Christopher; Montagu, Dominic; McFarland, Willi; Hlaing, Thaung; Khin, Hnin Su Su; San, Aung Kyaw; Briegleb, Christina; Chen, Ingrid; Sudhinaraset, May

    2015-03-06

    As efforts to contain artemisinin resistance and eliminate Plasmodium falciparum intensify, the accurate diagnosis and prompt effective treatment of malaria are increasingly needed in Myanmar and the Greater Mekong Sub-region (GMS). Rapid diagnostic tests (RDTs) have been shown to be safe, feasible, and effective at promoting appropriate treatment for suspected malaria, which are of particular importance to drug resistance containment. The informal private sector is often the first point of care for fever cases in malaria endemic areas across Myanmar and the GMS, but there is little published information about informal private provider practices, quality of service provision, or potential to contribute to malaria control and elimination efforts. This study tested different incentives to increase RDT use and improve the quality of care among informal private healthcare providers in Myanmar. The study randomized six townships in the Mon and Shan states of rural Myanmar into three intervention arms: 1) RDT price subsidies, 2) price subsidies with product-related financial incentives, and 3) price subsidies with intensified information, education and counselling (IEC). The study assessed the uptake of RDT use in the communities by cross-sectional surveys of 3,150 households at baseline and six months post-intervention (6,400 households total, 832 fever cases). The study also used mystery clients among 171 providers to assess quality of service provision across intervention arms. The pilot intervention trained over 600 informal private healthcare providers. The study found a price subsidy with intensified IEC, resulted in the highest uptake of RDTs in the community, as compared to subsidies alone or merchandise-related financial incentives. Moreover, intensified IEC led to improvements in the quality of care, with mystery client surveys showing almost double the number of correct treatment following diagnostic test results as compared to a simple subsidy. Results show

  1. Rapid and inexpensive body fluid identification by RNA profiling-based multiplex High Resolution Melt (HRM analysis [v1; ref status: indexed, http://f1000r.es/2hj

    Directory of Open Access Journals (Sweden)

    Erin K. Hanson

    2013-12-01

    Full Text Available Positive identification of the nature of biological material present on evidentiary items can be crucial for understanding the circumstances surrounding a crime. However, traditional protein-based methods do not permit the identification of all body fluids and tissues, and thus molecular based strategies for the conclusive identification of all forensically relevant biological fluids and tissues need to be developed. Messenger RNA (mRNA profiling is an example of such a molecular-based approach. Current mRNA body fluid identification assays involve capillary electrophoresis (CE or quantitative RT-PCR (qRT-PCR platforms, each with its own limitations. Both platforms require the use of expensive fluorescently labeled primers or probes. CE-based assays require separate amplification and detection steps thus increasing the analysis time. For qRT-PCR assays, only 3-4 markers can be included in a single reaction since each requires a different fluorescent dye. To simplify mRNA profiling assays, and reduce the time and cost of analysis, we have developed single- and multiplex body fluid High Resolution Melt (HRM assays for the identification of common forensically relevant biological fluids and tissues. The incorporated biomarkers include IL19 (vaginal secretions, IL1F7 (skin, ALAS2 (blood, MMP10 (menstrual blood, HTN3 (saliva and TGM4 (semen.  The HRM assays require only unlabeled PCR primers and a single saturating intercalating fluorescent dye (Eva Green. Each body-fluid-specific marker can easily be identified by the presence of a distinct melt peak. Usually, HRM assays are used to detect variants or isoforms for a single gene target. However, we have uniquely developed duplex and triplex HRM assays to permit the simultaneous detection of multiple targets per reaction. Here we describe the development and initial performance evaluation of the developed HRM assays. The results demonstrate the potential use of HRM assays for rapid, and relatively

  2. Rapid and inexpensive body fluid identification by RNA profiling-based multiplex High Resolution Melt (HRM analysis [v2; ref status: indexed, http://f1000r.es/314

    Directory of Open Access Journals (Sweden)

    Erin K. Hanson

    2014-02-01

    Full Text Available Positive identification of the nature of biological material present on evidentiary items can be crucial for understanding the circumstances surrounding a crime. However, traditional protein-based methods do not permit the identification of all body fluids and tissues, and thus molecular based strategies for the conclusive identification of all forensically relevant biological fluids and tissues need to be developed. Messenger RNA (mRNA profiling is an example of such a molecular-based approach. Current mRNA body fluid identification assays involve capillary electrophoresis (CE or quantitative RT-PCR (qRT-PCR platforms, each with its own limitations. Both platforms require the use of expensive fluorescently labeled primers or probes. CE-based assays require separate amplification and detection steps thus increasing the analysis time. For qRT-PCR assays, only 3-4 markers can be included in a single reaction since each requires a different fluorescent dye. To simplify mRNA profiling assays, and reduce the time and cost of analysis, we have developed single- and multiplex body fluid High Resolution Melt (HRM assays for the identification of common forensically relevant biological fluids and tissues. The incorporated biomarkers include IL19 (vaginal secretions, IL1F7 (skin, ALAS2 (blood, MMP10 (menstrual blood, HTN3 (saliva and TGM4 (semen.  The HRM assays require only unlabeled PCR primers and a single saturating intercalating fluorescent dye (Eva Green. Each body-fluid-specific marker can easily be identified by the presence of a distinct melt peak. Usually, HRM assays are used to detect variants or isoforms for a single gene target. However, we have uniquely developed duplex and triplex HRM assays to permit the simultaneous detection of multiple targets per reaction. Here we describe the development and initial performance evaluation of the developed HRM assays. The results demonstrate the potential use of HRM assays for rapid, and relatively

  3. 4. Payment Schemes

    Indian Academy of Sciences (India)

    Home; Journals; Resonance – Journal of Science Education; Volume 6; Issue 2. Electronic Commerce - Payment Schemes. V Rajaraman. Series Article Volume 6 Issue 2 February 2001 pp 6-13. Fulltext. Click here to view fulltext PDF. Permanent link: http://www.ias.ac.in/article/fulltext/reso/006/02/0006-0013 ...

  4. Alternative health insurance schemes

    DEFF Research Database (Denmark)

    Keiding, Hans; Hansen, Bodil O.

    2002-01-01

    In this paper, we present a simple model of health insurance with asymmetric information, where we compare two alternative ways of organizing the insurance market. Either as a competitive insurance market, where some risks remain uninsured, or as a compulsory scheme, where however, the level...... competitive insurance; this situation turns out to be at least as good as either of the alternatives...

  5. MIT Scheme Reference Manual

    Science.gov (United States)

    1991-01-01

    their Butterfly Scheme Reference, and to Margaret O’Connell for translating it from BBN’s text-formatting language to ours. Special thanks to Richard ... Stallman , Bob Chassell, and Brian Fox, all of the Free Software Foundation, for creating and maintaining the Texinfo formatting language in which this

  6. [Use of the rapid appraisal method for the identification of perceived needs in a low-class barrio in the city of Buenos Aires].

    Science.gov (United States)

    Lotersztain, M; Zorat, M; Lecouna, M; Motta, M

    2000-12-01

    To present rapid appraisal as a qualitative technique for researching the perceived needs of a population, which is valid, rapid and cheap. Qualitative method: semi-structured interviews with key informants, focus group. A poor quarter with 2921 inhabitants in the city of Buenos Aires. Key informants: people resident in the area for over 10 years. The main problems perceived by the community were: environmental pollution because of the accumulation of rubbish, alcohol and drug abuse among the young, and the need to extend the opening hours of the area's health centre. The needs perceived surpassed specific medical problems. They covered social questions, environmental problems and how to organise the health care system.

  7. A Scatter Storage Scheme for Dictionary Lookups

    Directory of Open Access Journals (Sweden)

    D. M. Murray

    1970-09-01

    Full Text Available Scatter storage schemes are examined with respect to their applicability to dictionary lookup procedures. Of particular interest are virtual scatter methods which combine the advantages of rapid search speed and reasonable storage requirements. The theoretical aspects of computing hash addresses are developed, and several algorithms are evaluated. Finally, experiments with an actual text lookup process are described, and a possible library application is discussed.

  8. Advances in rapid identification and susceptibility testing of bacteria in the clinical microbiology laboratory: implications for patient care and antimicrobial stewardship programs

    Directory of Open Access Journals (Sweden)

    Florian P. Maurer

    2017-03-01

    Full Text Available Early availability of information on bacterial pathogens and their antimicrobial susceptibility is of key importance for the management of infectious diseases patients. Currently, using traditional approaches, it usually takes at least 48 hours for identification and susceptibility testing of bacterial pathogens. Therefore, the slowness of diagnostic procedures drives prolongation of empiric, potentially inappropriate, antibacterial therapies. Over the last couple of years, the improvement of available techniques (e.g. for susceptibility testing, DNA amplification assays, and introduction of novel technologies (e.g. MALDI-TOF has fundamentally changed approaches towards pathogen identification and characterization. Importantly, these techniques offer increased diagnostic resolution while at the same time shorten the time-to-result, and are thus of obvious importance for antimicrobial stewardship. In this review, we will discuss recent advances in medical microbiology with special emphasis on the impact of novel techniques on antimicrobial stewardship programs.

  9. Advances in Rapid Identification and Susceptibility Testing of Bacteria in the Clinical Microbiology Laboratory: Implications for Patient Care and Antimicrobial Stewardship Programs.

    Science.gov (United States)

    Maurer, Florian P; Christner, Martin; Hentschke, Moritz; Rohde, Holger

    2017-03-30

    Early availability of information on bacterial pathogens and their antimicrobial susceptibility is of key importance for the management of infectious diseases patients. Currently, using traditional approaches, it usually takes at least 48 hours for identification and susceptibility testing of bacterial pathogens. Therefore, the slowness of diagnostic procedures drives prolongation of empiric, potentially inappropriate, antibacterial therapies. Over the last couple of years, the improvement of available techniques (e.g. for susceptibility testing, DNA amplification assays), and introduction of novel technologies (e.g. MALDI-TOF) has fundamentally changed approaches towards pathogen identification and characterization. Importantly, these techniques offer increased diagnostic resolution while at the same time shorten the time-to-result, and are thus of obvious importance for antimicrobial stewardship. In this review, we will discuss recent advances in medical microbiology with special emphasis on the impact of novel techniques on antimicrobial stewardship programs.

  10. Rapid Identification and Quantification of Natural Antioxidants in the Seeds of Rhubarb from Different Habitats in China Using Accelerated Solvent Extraction and HPLC-DAD-ESI-MSn-DPPH Assay.

    Science.gov (United States)

    Tan, Liang; Geng, Dan-dan; Hu, Feng-zu; Dong, Qi

    2016-01-01

    In this study, the 10 accessions of rhubarb seeds from different habitats in China were investigated. Lipids were removed using petroleum ether, and the effective components were then separated using accelerated solvent extraction with 80% aqueous methanol. An off-line 2,2-diphenyl-1-picrylhydrazyl (DPPH) free-radical scavenging method was used as the marker to evaluate the total antioxidant capability of extracts. On-line high-performance liquid chromatography-diode-array detectors-electrospray ionization-tandem mass spectrometry (HPLC-DAD-ESI-MS(n)) and HPLC-DAD-DPPH assays were developed for rapid identification and quantification of individual free-radical scavengers in extracts of rhubarb seeds. Ten free-radical scavengers from methanolic extracts of the rhubarb seeds were screened, five of which were identified and quantitatively analyzed: epicatechin, myricetin, hyperoside, quercitrin and quercetin. All were identified in rhubarb seeds for the first time and can be regarded as the major potent antioxidants in rhubarb seeds due to representing most of the total free-radical scavenging activity. Preliminary analysis of structures was performed for another five antioxidants. Based on our validation results, the developed method can be used for rapid separation, convenient identification and quantification of the multiple antioxidative constituents in rhubarb seeds, featuring good quantification parameters, accuracy and precision. The results are important to clarify the material basis and therapeutic mechanism of rhubarb seeds. © The Author 2015. Published by Oxford University Press. All rights reserved. For Permissions, please email: journals.permissions@oup.com.

  11. A Standardised Vocabulary for Identifying Benthic Biota and Substrata from Underwater Imagery: The CATAMI Classification Scheme.

    Directory of Open Access Journals (Sweden)

    Franziska Althaus

    Full Text Available Imagery collected by still and video cameras is an increasingly important tool for minimal impact, repeatable observations in the marine environment. Data generated from imagery includes identification, annotation and quantification of biological subjects and environmental features within an image. To be long-lived and useful beyond their project-specific initial purpose, and to maximize their utility across studies and disciplines, marine imagery data should use a standardised vocabulary of defined terms. This would enable the compilation of regional, national and/or global data sets from multiple sources, contributing to broad-scale management studies and development of automated annotation algorithms. The classification scheme developed under the Collaborative and Automated Tools for Analysis of Marine Imagery (CATAMI project provides such a vocabulary. The CATAMI classification scheme introduces Australian-wide acknowledged, standardised terminology for annotating benthic substrates and biota in marine imagery. It combines coarse-level taxonomy and morphology, and is a flexible, hierarchical classification that bridges the gap between habitat/biotope characterisation and taxonomy, acknowledging limitations when describing biological taxa through imagery. It is fully described, documented, and maintained through curated online databases, and can be applied across benthic image collection methods, annotation platforms and scoring methods. Following release in 2013, the CATAMI classification scheme was taken up by a wide variety of users, including government, academia and industry. This rapid acceptance highlights the scheme's utility and the potential to facilitate broad-scale multidisciplinary studies of marine ecosystems when applied globally. Here we present the CATAMI classification scheme, describe its conception and features, and discuss its utility and the opportunities as well as challenges arising from its use.

  12. Use of a Real-Time PCR TaqMan Assay for Rapid Identification and Differentiation of Burkholderia pseudomallei and Burkholderia mallei

    OpenAIRE

    U?Ren, Jana M.; Van Ert, Matthew N.; Schupp, James M.; Easterday, W Ryan; Simonson, Tatum S.; Okinaka, Richard T.; Pearson, Talima; Keim, Paul

    2005-01-01

    A TaqMan allelic-discrimination assay designed around a synonymous single-nucleotide polymorphism was used to genotype Burkholderia pseudomallei and Burkholderia mallei isolates. The assay rapidly identifies and discriminates between these two highly pathogenic bacteria and does not cross-react with genetic near neighbors, such as Burkholderia thailandensis and Burkholderia cepacia.

  13. Development of a rapid, cost-effective TaqMan Real-Time PCR Assay for identification and differentiation of Coccidioides immitis and Coccidioides posadasii.

    Science.gov (United States)

    Sheff, Kelly W; York, Emily R; Driebe, Elizabeth M; Barker, Bridget M; Rounsley, Steven D; Waddell, Victor G; Beckstrom-Sternberg, Stephen M; Beckstrom-Sternberg, James S; Keim, Paul S; Engelthaler, David M

    2010-05-01

    Coccidioidomycosis is an infection caused by Coccidioides immitis or C. posadasii. We developed a TaqMan real-time PCR assay that rapidly and accurately differentiates the species. This assay can be used as a tool to improve disease surveillance, increase understanding of the natural history of the infection, and assist in clinical differentiation studies.

  14. Prospective evaluation of a high multiplexing real-time polymerase chain reaction array for the rapid identification and characterization of bacteria causative of nosocomial pneumonia from clinical specimens: a proof-of-concept study.

    Science.gov (United States)

    Roisin, S; Huang, T-D; de Mendonça, R; Nonhoff, C; Bogaerts, P; Hites, M; Delaere, B; Hamels, S; de Longueville, F; Glupczynski, Y; Denis, O

    2017-09-27

    The purpose of this study was evaluation of the VAPChip assay based on the "Rapid-Array-PCR-technology" which targets 13 respiratory pathogens and 24 β-lactam resistance genes directly on respiratory clinical specimens. The first step included analysis of 45 respiratory specimens in order to calibrate and determine the threshold for target genes. The second prospective step involved 85 respiratory samples from patients suspected of nosocomial pneumonia collected in two academic hospitals over an 8-month period. Results of the VAPChip assay were compared to routine methods. The first step showed a large proportion of positive signals for H. influenzae and/or S. pneumoniae. For identification, discrepancies were observed in seven samples. Thresholds were adapted and two probes were re-designed to create a new version of the cartridge. In the second phase, sensitivity and specificity of the VAPchip for bacterial identification were 72.9% and 99.1%, respectively. Seventy (82%) pathogens were correctly identified by both methods. Nine pathogens detected by the VAPChip were culture negative and 26 pathogens identified by culture were VAPChip negative. For resistance mechanisms, 11 probes were positive without identification of pathogens with an antimicrobial-susceptibility testing compatible by culture. However, the patient's recent microbiological history was able to explain most of these positive signals. The VAPChip assay simultaneously detects different pathogens and resistance mechanisms directly from clinical samples. This system seems very promising but the extraction process needs to be automated for routine implementation. This kind of rapid point-of-care automated platform permitting a syndromic approach will be the future challenge in the management of infectious diseases.

  15. Selectively strippable paint schemes

    Science.gov (United States)

    Stein, R.; Thumm, D.; Blackford, Roger W.

    1993-03-01

    In order to meet the requirements of more environmentally acceptable paint stripping processes many different removal methods are under evaluation. These new processes can be divided into mechanical and chemical methods. ICI has developed a paint scheme with intermediate coat and fluid resistant polyurethane topcoat which can be stripped chemically in a short period of time with methylene chloride free and phenol free paint strippers.

  16. Signature Schemes Secure against Hard-to-Invert Leakage

    DEFF Research Database (Denmark)

    Faust, Sebastian; Hazay, Carmit; Nielsen, Jesper Buus

    2012-01-01

    -theoretically reveal the entire secret key. In this work, we propose the first constructions of digital signature schemes that are secure in the auxiliary input model. Our main contribution is a digital signature scheme that is secure against chosen message attacks when given an exponentially hard-to-invert function...... of the secret key. As a second contribution, we construct a signature scheme that achieves security for random messages assuming that the adversary is given a polynomial-time hard to invert function. Here, polynomial-hardness is required even when given the entire public-key – so called weak auxiliary input...... security. We show that such signature schemes readily give us auxiliary input secure identification schemes...

  17. The effect of rapid screening for methicillin-resistant Staphylococcus aureus (MRSA) on the identification and earlier isolation of MRSA-positive patients.

    LENUS (Irish Health Repository)

    Creamer, Eilish

    2010-04-01

    (1) To determine whether rapid screening with polymerase chain reaction (PCR) assays leads to the earlier isolation of patients at risk for methicillin-resistant Staphylococcus aureus (MRSA) colonization, (2) to assess compliance with routine MRSA screening protocols, (3) to confirm the diagnostic accuracy of the Xpert MRSA real-time PCR assay (Cepheid) by comparison with culture, and (4) to compare turnaround times for PCR assay results with those for culture results.

  18. A multiplex polymerase chain reaction based method for rapid identification of two species of the genus Scolytus Geoffroy (Col: Curculionidae: Scolytinae in Iran

    Directory of Open Access Journals (Sweden)

    S. Amini

    2016-04-01

    Full Text Available Molecular identification is going to be more widespread in taxonomic studies of insects when traditional tools are problematic and time consuming. Identification of bark beetles, as one of the most important pests of forests, based on morphological characteristics is difficult because of their small size and morphological similarities. In the current study, species-specific primers were desi gned to identify two most abundant and morphologically similar bark beetle species Scolytus ensifer Eichhoff 1881 and S. ecksteini Butovitsch 1929, both found on Ulmus minor Miller in north of Iran. These species-specific primers successfully produced a fragment size with 318 bp and 465 bp of mitochondrial cytochrome oxidase 1 (CO1 gene in S. ensifer and S. ecksteini respectively. The results revealed tha t the multiplex polymerase chain reaction using the species-specific primers could amplify a unique band to distinguish these two species so confirmed this method as a convenient and quick tool to identify those two bark beetle species.

  19. Rapid identification of pathogens directly from blood culture bottles by Bruker matrix-assisted laser desorption laser ionization-time of flight mass spectrometry versus routine methods.

    Science.gov (United States)

    Jamal, Wafaa; Saleem, Rola; Rotimi, Vincent O

    2013-08-01

    The use of matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF MS) for identification of microorganisms directly from blood culture is an exciting dimension to the microbiologists. We evaluated the performance of Bruker SepsiTyper kit™ (STK) for direct identification of bacteria from positive blood culture. This was done in parallel with conventional methods. Nonrepetitive positive blood cultures from 160 consecutive patients were prospectively evaluated by both methods. Of 160 positive blood cultures, the STK identified 114 (75.6%) isolates and routine conventional method 150 (93%). Thirty-six isolates were misidentified or not identified by the kit. Of these, 5 had score of >2.000 and 31 had an unreliable low score of time using the STK was 35 min, including extraction steps and 30:12 to 36:12 h with routine method. The STK holds promise for timely management of bacteremic patients. Copyright © 2013 Elsevier Inc. All rights reserved.

  20. CANONICAL BACKWARD DIFFERENTIATION SCHEMES FOR ...

    African Journals Online (AJOL)

    CANONICAL BACKWARD DIFFERENTIATION SCHEMES FOR SOLUTION OF NONLINEAR INITIAL VALUE PROBLEMS OF FIRST ORDER ORDINARY DIFFERENTIAL EQUATIONS. ... Global Journal of Mathematical Sciences ... KEY WORDS: backward differentiation scheme, collocation, initial value problems. Global Jnl ...

  1. Low-pass shotgun sequencing of the barley genome facilitates rapid identification of genes, conserved non-coding sequences and novel repeats

    Directory of Open Access Journals (Sweden)

    Graner Andreas

    2008-10-01

    Full Text Available Abstract Background Barley has one of the largest and most complex genomes of all economically important food crops. The rise of new short read sequencing technologies such as Illumina/Solexa permits such large genomes to be effectively sampled at relatively low cost. Based on the corresponding sequence reads a Mathematically Defined Repeat (MDR index can be generated to map repetitive regions in genomic sequences. Results We have generated 574 Mbp of Illumina/Solexa sequences from barley total genomic DNA, representing about 10% of a genome equivalent. From these sequences we generated an MDR index which was then used to identify and mark repetitive regions in the barley genome. Comparison of the MDR plots with expert repeat annotation drawing on the information already available for known repetitive elements revealed a significant correspondence between the two methods. MDR-based annotation allowed for the identification of dozens of novel repeat sequences, though, which were not recognised by hand-annotation. The MDR data was also used to identify gene-containing regions by masking of repetitive sequences in eight de-novo sequenced bacterial artificial chromosome (BAC clones. For half of the identified candidate gene islands indeed gene sequences could be identified. MDR data were only of limited use, when mapped on genomic sequences from the closely related species Triticum monococcum as only a fraction of the repetitive sequences was recognised. Conclusion An MDR index for barley, which was obtained by whole-genome Illumina/Solexa sequencing, proved as efficient in repeat identification as manual expert annotation. Circumventing the labour-intensive step of producing a specific repeat library for expert annotation, an MDR index provides an elegant and efficient resource for the identification of repetitive and low-copy (i.e. potentially gene-containing sequences regions in uncharacterised genomic sequences. The restriction that a particular

  2. Bonus Schemes and Trading Activity

    NARCIS (Netherlands)

    Pikulina, E.S.; Renneboog, L.D.R.; Ter Horst, J.R.; Tobler, P.N.

    2013-01-01

    Abstract: Little is known about how different bonus schemes affect traders’ propensity to trade and which bonus schemes improve traders’ performance. We study the effects of linear versus threshold (convex) bonus schemes on traders’ behavior. Traders purchase and sell shares in an experimental stock

  3. Bonus schemes and trading activity

    NARCIS (Netherlands)

    Pikulina, E.S.; Renneboog, L.D.R.; ter Horst, J.R.; Tobler, P.N.

    2014-01-01

    Little is known about how different bonus schemes affect traders' propensity to trade and which bonus schemes improve traders' performance. We study the effects of linear versus threshold bonus schemes on traders' behavior. Traders buy and sell shares in an experimental stock market on the basis of

  4. [Rapid screening and identification of 22 allergenic disperse dyes in ecological textiles by high performance liquid chromatography-linear ion trap/orbitrap mass spectrometry].

    Science.gov (United States)

    Niu, Zengyuan; Luo, Xin; Ye, Xiwen; Xiu, Xiaoli; Zhang, Li; Wang, Xin; Chen, Jing

    2015-10-01

    A rapid screening method based on high performance liquid chromatography-linear ion trap/orbitrap high-resolution mass spectrometry (HPLC-LTQ/Orbitrap MS) for 22 disperse dyes in ecological textiles has been established. The target compounds were extracted by pyridine/water (1:1, v/v) by shaking extraction in 90 degrees C water bath. The extracts were then separated by a CAPCELL PAK C18 column (100 mm x 2.0 mm, 5 μm) using gradient elution with acetonitrile-5 mmol/L ammonium acetate containing 0.01% (v/v) formic acid as mobile phases, and finally analyzed by HPLC-LTQ/Orbitrap in positive and negative ESI modes. The retention time and accurate mass of parent ion were used for fast screening of 22 disperse dyes, while the confirmatory analysis was obtained by fragments generated by collision-induced dissociation (CID) MS/MS. Target analysis exhibited high mass accuracy (textiles, and Disperse Orange 37/76 was detected in one of them. With high selectivity and strong anti-jamming ability, this method is simple, rapid, accurate, and it can be used for the inspection of disperse dyes in textiles.

  5. Assessing direct analysis in real-time-mass spectrometry (DART-MS) for the rapid identification of additives in food packaging.

    Science.gov (United States)

    Ackerman, L K; Noonan, G O; Begley, T H

    2009-12-01

    The ambient ionization technique direct analysis in real time (DART) was characterized and evaluated for the screening of food packaging for the presence of packaging additives using a benchtop mass spectrometer (MS). Approximate optimum conditions were determined for 13 common food-packaging additives, including plasticizers, anti-oxidants, colorants, grease-proofers, and ultraviolet light stabilizers. Method sensitivity and linearity were evaluated using solutions and characterized polymer samples. Additionally, the response of a model additive (di-ethyl-hexyl-phthalate) was examined across a range of sample positions, DART, and MS conditions (temperature, voltage and helium flow). Under optimal conditions, molecular ion (M+H+) was the major ion for most additives. Additive responses were highly sensitive to sample and DART source orientation, as well as to DART flow rates, temperatures, and MS inlet voltages, respectively. DART-MS response was neither consistently linear nor quantitative in this setting, and sensitivity varied by additive. All additives studied were rapidly identified in multiple food-packaging materials by DART-MS/MS, suggesting this technique can be used to screen food packaging rapidly. However, method sensitivity and quantitation requires further study and improvement.

  6. A multiple-dimension liquid chromatography coupled with mass spectrometry data strategy for the rapid discovery and identification of unknown compounds from a Chinese herbal formula (Er-xian decoction).

    Science.gov (United States)

    Wang, Caihong; Zhang, Jinlan; Wu, Caisheng; Wang, Zhe

    2017-10-06

    It is very important to rapidly discover and identify the multiple components of traditional Chinese medicine (TCM) formula. High performance liquid chromatography with high resolution tandem mass spectrometry (HPLC-HRMS/MS) has been widely used to analyze TCM formula and contains multiple-dimension data including retention time (RT), high resolution mass (HRMS), multiple-stage mass spectrometric (MSn), and isotope intensity distribution (IID) data. So it is very necessary to exploit a useful strategy to utilize multiple-dimension data to rapidly probe structural information and identify chemical compounds. In this study, a new strategy to initiatively use the multiple-dimension LC-MS data has been developed to discover and identify unknown compounds of TCM in many styles. The strategy guarantees the fast discovery of candidate structural information and provides efficient structure clues for identification. The strategy contains four steps in sequence: (1) to discover potential compounds and obtain sub-structure information by the mass spectral tree similarity filter (MTSF) technique, based on HRMS and MSn data; (2) to classify potential compounds into known chemical classes by discriminant analysis (DA) on the basis of RT and HRMS data; (3) to hit the candidate structural information of compounds by intersection sub-structure between MTSF and DA (M,D-INSS); (4) to annotate and confirm candidate structures by IID data. This strategy allowed for the high exclusion efficiency (greater than 41%) of irrelevant ions in er-xian decoction (EXD) while providing accurate structural information of 553 potential compounds and identifying 66 candidates, therefore accelerating and simplifying the discovery and identification of unknown compounds in TCM formula. Copyright © 2017 Elsevier B.V. All rights reserved.

  7. The identification of fall history using maximal and rapid isometric torque characteristics of the hip extensors in healthy, recreationally active elderly females: a preliminary investigation.

    Science.gov (United States)

    Palmer, Ty B; Thiele, Ryan M; Williams, Katherine B; Adams, Bailey M; Akehi, Kazuma; Smith, Douglas B; Thompson, Brennan J

    2015-08-01

    Maximal and rapid torque characteristics of the hip extensor muscles play an important role in fall prevention and other balance-related performances; however, few studies have investigated the ability of these variables at identifying fall-history status in healthy, recreationally active elderly adults. This study aimed to examine the effectiveness of maximal and rapid isometric torque characteristics of the hip extensor muscles to differentiate between healthy, recreationally active elderly females with (fallers) and without (non-fallers) a history a falls. Six elderly female fallers (mean ± SD: age = 73 ± 7 year; mass = 68 ± 16 kg; height = 160 ± 5 cm) and nine elderly female non-fallers (age = 71 ± 7 year; mass = 66 ± 16 kg; height = 157 ± 6 cm) performed two isometric maximal voluntary contractions (MVCs) of the hip extensor muscles. Peak torque (PT) and absolute and relative rate of torque development (RTD) at the early (0-50 ms) and late (100-200 ms) phases of muscle contraction were examined during each MVC. Absolute and relative RTD at 0-50 ms were greater (P = 0.039 and 0.011, respectively) in the non-fallers compared to the fallers. However, no group-related differences (P = 0.160-0.573) were observed for PT nor absolute and relative RTD at 100-200 ms. Early rapid strength production of the hip extensor muscles may be a sensitive and effective measure for discriminating between elderly females of different fall histories. These findings may provide important insight regarding implications for the assessment of fall risk and in the development of proper training programs aimed at minimizing the occurrence of falls and other balance-related injuries in the elderly.

  8. Analytical strategy for rapid identification and quantification of lubricant additives in mineral oil by high-performance thin-layer chromatography with UV absorption and fluorescence detection combined with mass spectrometry and infrared spectroscopy.

    Science.gov (United States)

    Dytkiewitz, Elisabeth; Morlock, Gertrud E

    2008-01-01

    A simple strategy for identification and quantification of lubricant additives in mineral oil was demonstrated by high-performance thin-layer chromatography with UV absorption and fluorescence detection using various coupling options, e.g., with attenuated total reflectance infrared (ATR-IR) spectroscopy, Fourier transform infrared (FTIR) spectroscopy, electrospray ionization mass spectrometry (ESI-MS), and direct analysis in real-time mass spectrometry (DART-MS). For the additives zinc bis(O,O'-diisobutyl dithiophosphate), zinc bis(O,O'-didodecyl dithiophosphate), and Anglamol 99, 2 chromatographic systems were developed, i.e., a reversed-phase (RP) system on RP2 plates using an acetonitrile-based mobile phase and a normal-phase system on silica gel 60 plates using a toluene-based gradient. Densitometry was performed by absorption measurement at 220 nm. Repeatabilities (relative standard deviation, n = 6) between 2.2 and 5.5% and correlation coefficients >0.9973 were highly satisfactory for the analysis of these additives in the mineral oil. Primuline reagent was used to improve the detection limit of the lipophilic additives by a factor of 2, followed by fluorescence measurement at UV 366/>400 nm. For rapid identification by ATR-IR and FTIR, the respective additive zones on the plate were online extracted by an interface called ChromeXtract, concentrated, and directly applied for measurements in the wave number range of 4000-400 cm(-1). Identification was confirmed by online ESI-MS within a minute using ChromeXtract and by DART-MS within seconds.

  9. Rapid characterisation and identification of compounds in Saposhnikoviae Radix by high-performance liquid chromatography coupled with electrospray ionisation quadrupole time-of-flight mass spectrometry.

    Science.gov (United States)

    Chen, Luxiao; Chen, Xiangyang; Su, Lei; Jiang, Yanyan; Liu, Bin

    2017-08-18

    Saposhnikoviae Radix (SR), the dried root of Saposhnikovia divaricata (Turcz.) Schischk. (Umbelliferae), is commonly used as a traditional Chinese medicine. In this study, a rapid and accurate method was firstly, developed for the qualitative analysis of SR by high-performance liquid chromatography coupled with electrospray ioni