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Results for “microbial detection”

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At least 163 records · Page 9Linked to original sources

PubMed2004

Microbial recognition by Toll-like receptors.

Toll-like receptors (TLRs) sense invasion of microorganisms by detecting microbial components that are conserved among pathogens. Recognition of microbial components by TLRs triggers activation of the innate immune system. Signaling pathways via TLRs originate from conserved cytoplasmic Toll/IL-1 receptor (TIR) domains. Recent accumulating evidence demonstrates that TIR domain-containing adaptors, such as MyD88, TIRAP/Mal, TRIF, and TRAM, regulate TLR-mediated signaling pathways. MyD88 is common to all TLR-mediated pathways, which lead to the production of inflammatory cytokines, whereas TRIF mediates induction of IFN-beta in TLR3 and TLR4 signaling pathways. TIRAP/Mal is implicated in the TLR2- and TLR4-mediated MyD88-dependent signaling pathway. TRAM is specifically involved in the TLR4-mediated TRIF-dependent pathway. Thus, TIR domain-containing adaptors play a pivotal role in TLR signaling pathways, which culminate in pathogen-specific immune responses.

Adaptor Proteins, Signal Transducing↗

PubMed1983

Nonculture techniques using blood specimens for the diagnosis of infectious disease.

Recent technological advances have increased the use of blood as a source for diagnosis of local and systemic infectious diseases. Traditionally, viable organisms may be cultured from the blood and others identified microscopically. The ability to detect microbial products by physicochemical, bioactivity, and antigenic techniques and the ability to detect specific immunoglobulin M (IgM) responses to infection improve our diagnostic capabilities but also present new problems in determining clinical relevance. The diagnostic use and reproducibility of new tests should be verified by comparative clinical field testing before implementation.

Antigen-Antibody Complex↗

PubMed2004

Filter-based microfluidic device as a platform for immunofluorescent assay of microbial cells.

A filter-based microfluidic device was combined with immunofluorescent labeling as a platform to rapidly detect microbial cells. The coin-sized device consisted of micro-chambers, micro-channels and filter weirs (gap = 1-2 microm), and was demonstrated to effectively trap and concentrate microbial cells (i.e., Cryptosporidium parvum and Giardia lamblia), which were larger in size than the weir gap. After sample injection, a staining solution containing fluorescently-labeled antibodies was continuously provided into the device (flow rate = 20 microl min(-1)) to flush the microbial cells toward the weirs and to accelerate the fluorescent labeling reaction. Using a staining solution that was 10 to 100 times more dilute than the recommended concentration used in a conventional glass method, those target cells with a fluorescent signal-to-noise ratio of 12 could be microscopically observed at single-cell level within 2 to 5 min prior to secondary washing.

Animals↗

PubMed1993

Diagnostic microbiology of bacterial vaginosis.

Bacterial vaginosis is characterized microbiologically by replacement of the Lactobacillus-predominant vaginal flora by Gardnerella vaginalis, Bacteroides species, Mobiluncus species, and genital mycoplasmas. A standardized, laboratory-based diagnostic test for bacterial vaginosis is desirable in those instances in which a microscope is unavailable in the clinic or when the evaluator of the wet mount examination is inexperienced at recognizing clue cells. Vaginal cultures have excellent sensitivity for the diagnosis of bacterial vaginosis, but because the predictive value of a positive G. vaginalis culture is less than 50%, cultures are not recommended. Vaginal Gram smears are objective and reproducible, have 62% to 100% sensitivity, and have a positive predictive value of 76% to 100%. Diagnosis of bacterial vaginosis with use of Papanicolaou-stained smears has been reported, but standardized criteria have not been widely adopted. A rapid, office-based oligonucleotide probe test for high concentrations of G. vaginalis has been developed and may be useful when direct microscopy is unavailable or confidence in microscopic examination is low. Laboratory tests that detect microbial products unique to the vaginal fluid of women with bacterial vaginosis include detection of amines (putrescine, cadaverine, and trimethylamine), measurement of the relative levels of succinate and lactate in the vaginal fluid by gas chromatography, detection of proline aminopeptidase by colorimetric assay, or detection of sialidases in the vaginal fluid. Although these tests are not yet applicable to routine use, these research tests could be adapted for wider use in office laboratories. Vaginal Gram stains are more useful than culture for laboratory confirmation of bacterial vaginosis.

Bacteriological Techniques↗

PubMed2026

Oral cavity as a reservoir of respiratory pathogens: microbial dynamics in patients receiving mechanical ventilation.

OBJECTIVE: To investigate the prevalence, co-occurrence, and temporal dynamics of Staphylococcus aureus, Klebsiella pneumoniae, Streptococcus pneumoniae, and Pseudomonas aeruginosa in oral and tracheal samples from critically ill patients receiving mechanical ventilation. METHODS: This observational analytical study included 21 adult patients receiving mechanical ventilation who were admitted to an intensive care unit. Oral and tracheal samples were collected within the first 24 h after intubation (Day 0) and again after 48 h (Day 2). Bacterial genomic deoxyribonucleic acid was extracted and analyzed using quantitative realtime polymerase chain reaction. Microbial detection was evaluated qualitatively based on the presence or absence of each microorganism and quantitatively using relative bacterial load analysis. Statistical analyses assessed correlations between oral and tracheal colonization patterns and temporal changes in bacterial load. RESULTS: More than 43% of oral and tracheal samples tested positive for at least one investigated pathogen at both collection time points, indicating persistent microbial colonization during mechanical ventilation. Significant positive correlations for Staphylococcus aureus detection were observed between oral and tracheal samples on Day 0 (p=0.0028) and Day 2 (p=0.0021), suggesting a possible association between oral colonization and lower airway microbial presence. Streptococcus pneumoniae detected in initial oral samples showed a significant correlation with tracheal detection after 48 h (p=0.03). Quantitative analyses demonstrated no significant changes in bacterial load over time for most microorganisms. However, Klebsiella pneumoniae showed a significant increase in tracheal bacterial load between Day 0 and Day 2 (p=0.04), with higher tracheal loads than oral samples on Day 2 (p=0.01). CONCLUSION: The oral cavity may serve as an important reservoir of respiratory pathogens in patients receiving mechanical ventilation. The observed correlations between oral and tracheal colonization and pathogen-specific colonization patterns reinforce the relevance of oral health management in critical care settings. In particular, the increase in tracheal Klebsiella pneumoniae load suggests that lower airway proliferation may occur during mechanical ventilation independently of simultaneous increases in oral bacterial load.

Humans↗

PubMed2006

Real-time fluorogenic reverse transcription-PCR assays for detection of bacteriophage MS2.

Bacteriophage MS2 is used in place of pathogenic viruses in a wide variety of studies that range from testing of compounds for disinfecting surfaces to studying environmental transport and fate of pathogenic viruses in groundwater. MS2 is also used as a pathogen simulant in the research, development, and testing (including open air tests) of methods, systems, and devices for the detection of pathogens in both the battlefield and homeland defense settings. PCR is often used as either an integral part of such detection systems or as a reference method to assess the sensitivity and specificity of microbial detection. To facilitate the detection of MS2 by PCR, we describe here a set of real-time fluorogenic reverse transcription-PCR assays. The sensitivity of the assays (performed with primer pairs and corresponding dye-labeled probes) ranged from 0.4 to 40 fg of MS2 genomic RNA (200 to 20,000 genome equivalents). We also demonstrate the usefulness of the primer pairs in assays without dye-labeled probe that included the DNA-binding dye SYBR green. None of the assays gave false-positive results when tested against 400 pg of several non-MS2 nucleic acid targets.

DNA Primers↗

PubMed1998

Polymerase chain reaction in the detection of microbes in amniotic fluid.

Intra-amniotic infection during pregnancy can be caused by bacteria, viruses or protozoa, Toxoplasma gondii for example. Bacterial intrauterine infections are connected with premature birth, premature rupture of fetal membranes, and infective complications of both the mother and the newborn. Viral infections and Toxoplasma gondii can cause fetal malformations and illness with serious sequelae to the infant or fetal death in utero. Determining the causative agent is important and often greatly affects the prognosis of the newborn. Amniotic fluid is in most cases easily and safely obtainable during the second and third trimester and can be used in several microbiological assays. These include bacterial and viral cultures, Gram staining, quantitative assays for immunoglobulins or cytokines, and polymerase chain reaction (PCR) for detecting microbial DNA. This review concentrates on broad-spectrum or universal bacterial PCR for detection of bacterial DNA in amniotic fluid and on PCR assays for certain clinically important viruses and for Toxoplasma gondii.

Amniotic Fluid↗

PubMed1999

[Relationships between air conditioning, airborne microorganisms and health].

Concurrently with the increase of air-conditioning, potentially severe or frequent new diseases have emerged, giving rise to social and economical consequences. The first part of this work is a state of the art review of the relationships between air-conditioning, airborne microorganisms and health, through a technical, metrological and medical approach. The second part presents four studies performed in this field. Two of them deal with the relationship between airborne microorganisms and technical features of air-conditioning. Measurements performed on actual sites demonstrated the benefit of using high efficiency filters and low risk components in air-conditioning systems. The third study was aimed to look for a relationship between airborne microorganisms and sick building syndrome symptoms. Statistical analyses of individual data revealed significant associations between airborne bacteria or fungi and symptoms. These results may be the first step in determining a dose-response relationship, in order to define threshold limit values in this field. In the fourth study, the contribution of particle counting in assessing exposure to airborne microorganisms was explored by monitoring simultaneous variations of microbial and particle concentrations. The results showed that associating particle counting may allow to detect microbial variations instantaneously, and therefore improve the assessment of exposure to airborne microorganisms.

Air Conditioning↗

PubMed2003

Detection of cyclic lipopeptide biomarkers from Bacillus species using atmospheric pressure matrix-assisted laser desorption/ionization mass spectrometry.

A novel approach to microbial detection using atmospheric pressure matrix-assisted laser desorption/ionization with an ion trap mass spectrometer to analyze whole cell bacteria is introduced. This new approach was tested with lyophilized spores and cultures of Bacillus globigii (BG) grown on agar media for 4 days or longer. At each stage of growth, it was found that biomarkers, identified as cyclic lipopeptides known as fengycin and surfactin, could be detected by pulsed ultraviolet laser irradiation of intact BG cells (approximately 5 mg) cocrystallized with alpha-cyano-4-hydroxycinnamic acid. Furthermore, definitive amino acid sequence information was obtained by performing tandem mass spectrometry on the precursor ions of the cyclic lipopeptides. The investigation was broadened to include the examination of aerosolized BG spores collected from the atmosphere and directly deposited onto double-sided tape. Subsequent analysis of the recovered spores resulted in the production of mass peaks consistent with fengycin. Other Bacillus species were analyzed for comparison and showed mass spectral peaks also identified as originating from various cyclic lipopeptides. Further studies were conducted using a pulsed infrared laser as the excitation source to analyze BG cells (approximately 5 mg) suspended in a matrix of 0.03 M ammonium citrate and glycerol resulting in the production of ions characteristic of fengycin and surfactin.

Aerosols↗

PubMed2005

Use of a solid-phase fluorescent cytometric technique for the detection of bacteria in platelet concentrates.

Blood services worldwide are now striving to reduce the risk of transmission of bacteria by transfusion. The BacT/ALERT microbial detection system (bioMerieux, Basingstoke, Hants, UK) is currently regarded as the 'gold standard' for bacterial screening of platelet concentrates. The BacT/ALERT is a culture system and will not generate an 'instant' (within 2 h) determination. We report on the Scansystem (Hemosystem, Marseille, France), a solid-phase fluorescent cytometric technique, which enables the rapid detection of bacteria (within 90 min) in platelet concentrates. The study was performed in two parts - one involving the routine screening of platelet concentrates and the other determining the sensitivity of the system. In both arms of the study, the BacT/ALERT was used for comparative purposes. In total, 900 platelet concentrates were screened (63 apheresis and 837 buffy coat pooled). No bacteria were detected in any of the platelet concentrates tested by means of either the Scansystem or the BacT/ALERT. The sensitivity of the Scansystem was in the order of 10(3) cfu mL(-1). Escherichia coli and Staphylococcus aureus were detected by using the Scansystem at 1 cfu mL(-1). The BacT/ALERT detected all organisms tested (n = 6) at 1 cfu mL(-1). The Scansystem offers a sensitive alternative technology to bacterial culture, with the benefit of a rapid test time.

Bacteria↗

PubMed1990

Aladapcin, a new microbial metabolite that enhances host resistance against bacterial infection. Production, isolation, physico-chemical properties and biological activities.

We have constructed a new screening system for detecting microbial products that enhance host resistance against bacterial infection. It was found that a new compound with such activity is produced by a soil isolate classified as Nocardia sp. SANK 60484. The compound was isolated from the culture filtrate of the organism and named aladapcin after its amino acid composition. Aladapcin was obtained as an amphoteric white amorphous powder with the molecular formula, C13H25N5O5. It consists of 2 mol of D-alanine and 1 mol of meso-diaminopimelic acid. From the analysis of IR, 1H NMR and FAB-MS spectra, the structure was assigned to be a tripeptide. Aladapcin enhanced host resistance against an experimental Escherichia coli infection in mice at doses ranging between 1 and 100 micrograms/kg.

Adjuvants, Immunologic↗

PubMed2005

Applications of Luminex xMAP technology for rapid, high-throughput multiplexed nucleic acid detection.

BACKGROUND: As we enter the post-genome sequencing era and begin to sift through the enormous amount of genetic information now available, the need for technologies that allow rapid, cost-effective, high-throughput detection of specific nucleic acid sequences becomes apparent. Multiplexing technologies, which allow for simultaneous detection of multiple nucleic acid sequences in a single reaction, can greatly reduce the time, cost and labor associated with single reaction detection technologies. METHODS: The Luminex xMAP system is a multiplexed microsphere-based suspension array platform capable of analyzing and reporting up to 100 different reactions in a single reaction vessel. This technology provides a new platform for high-throughput nucleic acid detection and is being utilized with increasing frequency. Here we review specific applications of xMAP technology for nucleic acid detection in the areas of single nucleotide polymorphism (SNP) genotyping, genetic disease screening, gene expression profiling, HLA DNA typing and microbial detection. CONCLUSIONS: These studies demonstrate the speed, efficiency and utility of xMAP technology for simultaneous, rapid, sensitive and specific nucleic acid detection, and its capability to meet the current and future requirements of the molecular laboratory for high-throughput nucleic acid detection.

Animals↗

PubMedDate not supplied

Molecular diagnosis of microbial contamination in cosmetic and pharmaceutical products: a review.

Molecular methodologies such as adenosine triphosphate (ATP) bioluminescence and polymerase chain reaction (PCR)-based assays provide rapid quality control analysis of cosmetic and pharmaceutical finished products and raw materials. Using a single enrichment broth for bacteria, yeast, and mold, ATP bioluminescence detected microbial contamination within 27 h. Samples were automatically lysed to release microbial ATP and light production was quantitated using the Celsis Optocomp. However, to maintain the detection time to within 27 h, different enrichment broths were required for neutralization of antimicrobial ingredients in finished products and to provide specific nutrients for growth optimization. To perform the PCR reaction, bacterial DNA was extracted using a Tris-EDTA-Tween 20-proteinase K buffer at 35 degrees C while yeast and mold DNA were extracted using a Tris-EDTA-SDS buffer at 95 degrees C. Extracted microbial DNA was added to Ready-To-Go PCR beads and specific DNA primers. The primers were targeted to amplify specific regions within Escherichia coli, Pseudomonas aeruginosa, Staphylococcus aureus, Salmonella typhimurium, Burkholderia cepacia, Candida albicans, and Aspergillus niger. Furthermore, conserved bacterial ribosomal DNA sequences have also been used for sterility testing of samples. The results from these studies indicate that both ATP bioluminescence and PCR assays provide rapid, reliable, and cost effective methods for quality evaluation. This will ultimately result in faster product release and production optimization.

Adenosine Triphosphate↗

PubMed2004

Detection of low levels of Listeria monocytogenes cells by using a fiber-optic immunosensor.

Biosensor technology has a great potential to meet the need for sensitive and nearly real-time microbial detection from foods. An antibody-based fiber-optic biosensor to detect low levels of Listeria monocytogenes cells following an enrichment step was developed. The principle of the sensor is a sandwich immunoassay where a rabbit polyclonal antibody was first immobilized on polystyrene fiber waveguides through a biotin-streptavidin reaction to capture Listeria cells on the fiber. Capture of cells on the fibers was confirmed by scanning electron microscopy. A cyanine 5-labeled murine monoclonal antibody, C11E9, was used to generate a specific fluorescent signal, which was acquired by launching a 635-nm laser light from an Analyte 2000 and collected by a photodetector at 670 to 710 nm. This immunosensor was specific for L. monocytogenes and showed a significantly higher signal strength than for other Listeria species or other microorganisms, including Escherichia coli, Enterococcus faecalis, Salmonella enterica, Lactobacillus plantarum, Carnobacterium gallinarum, Hafnia alvei, Corynebacterium glutamicum, Enterobacter aerogenes, Pseudomonas aeruginosa, and Serratia marcescens, in pure or in mixed-culture setup. Fiber-optic results could be obtained within 2.5 h of sampling. The sensitivity threshold was about 4.3 x 10(3) CFU/ml for a pure culture of L. monocytogenes grown at 37 degrees C. When L. monocytogenes was mixed with lactic acid bacteria or grown at 10 degrees C with 3.5% NaCl, the detection threshold was 4.1 x 10(4) or 2.8 x 10(7) CFU/ml, respectively. In less than 24 h, this method could detect L. monocytogenes in hot dog or bologna naturally contaminated or artificially inoculated with 10 to 1,000 CFU/g after enrichment in buffered Listeria enrichment broth.

Animals↗

PubMed1996

Enzyme-linked immunomagnetic electrochemical detection of Salmonella typhimurium.

There is a need for rapid methods to detect pathogenic bacteria in food products as alternatives to the current laborious and time-consuming culture procedures. We report a microbial detection technique that combines the selectivity of antibody-coated superparamagnetic beads with the rapidity and sensitivity of electrochemical detection in a format termed enzyme-linked immunomagnetic electrochemistry. In it, Salmonella typhimurium were sandwiched between antibody-coated magnetic beads and an enzyme-conjugated antibody. With the aid of a magnet, the beads (with or without bound bacteria) were localized onto the surface of disposable graphite ink electrodes in a multi-well plate format. Enzyme substrate was added and conversion of substrate to an electroactive product was measured using electrochemical detection. The electrochemical response was directly proportional to the number of captured bacteria. Using this technique, a minimum detectable level of 8 x 10(3) cells/ml of Salmonella typhimurium in buffer was achieved in ca. 80 min.

Electrochemistry↗

PubMed2004

Innate immunity via Toll-like receptors and Nod proteins.

Host defense against microbes requires the development of an efficient immune response aimed to eradicate the source of infection. Through the expression of a battery of germ-line encoded receptors, including the Toll-like receptors and Nod proteins, the innate immune system, which is a prerequisite to the adaptive immune response, detects microbial motifs and initiates pro-inflammatory signaling. Current research into innate immune function focuses on the nature of the ligands detected by this system, the cell signaling that occurs downstream of receptor activation and finally, how these signals culminate into a tailored adaptive immune response directed to eradicate a specific infection.

Animals↗

PubMed1995

Detection of Bartonella (Rochalimaea) henselae bacteremia using BacT/Alert blood culture system.

Bartonella henselae was isolated from the blood of five febrile immunosuppressed patients using BacT/Alert (Organon Teknika, Durham, NC) automated microbial detection system. An immunofluorescence assay (using 1:1000 dilutions) was used to confirm identification of fastidious, pleomorphic, non-Gram staining, argyrophilic bacilli displaying rachety motility that had been presumptively identified as Bartonella spp. The practicality of identification of Bartonella henselae using goat antisera for use in a routine clinical microbiology laboratory was demonstrated by this study.

AIDS-Related Opportunistic Infections↗

PubMed2005

Layered peptide arrays: high-throughput antibody screening of clinical samples.

High-throughput methods to detect and quantify antibodies in sera and other patient specimens have use for many clinical and laboratory studies, including those associated with cancer detection, microbial exposures, and autoimmune diseases. We developed a new technique, termed layered peptide array (LPA), to serve as a screening tool to detect antibodies in a highly multiplexed format. We demonstrate here that a prototype LPA was capable of producing approximately 5000 measurements per experiment and appeared to be scalable to higher throughput levels. Sera and saliva from Sjögren's syndrome patients served as a test set to examine antibody titers in clinical samples. The LPA platform exhibited both a high sensitivity (100%) and high specificity (94%) for correctly identifying SSB antigen-positive samples. The multiplex capability of the platform was also confirmed when serum and saliva samples were analyzed for antibody reactivity to several peptides, including Sjögren's syndrome antigens A and B. The data indicate that LPA analysis will be a useful method for a number of screening applications.

Antibodies↗