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Bacterial Concentrations (bacterial + concentration)
Selected AbstractsDirect detection of bacterial pathogens in representative dairy products using a combined bacterial concentration-PCR approachJOURNAL OF APPLIED MICROBIOLOGY, Issue 6 2004K.A. Stevens Abstract Aims:, To develop a simple, rapid method to concentrate and purify bacteria and their nucleic acids from complex dairy food matrices in preparation for direct pathogen detection using polymerase chain reaction (PCR). Methods and Results:, Plain non-fat yogurt and cheddar cheese were each seeded with Listeria monocytogenes or Salmonella enterica serovar. Enteritidis in the range of 101,106 CFU per 11-g sample. Samples were then processed for bacterial concentration using high-speed centrifugation (9700 g) followed by DNA extraction, PCR amplification, and amplicon confirmation by hybridization. Bacterial recoveries after centrifugation ranged from 53 to >100% and 71 to >100% for serovar. Enteritidis and L. monocytogenes, respectively, in the non-fat yogurt samples; and from 77 to >100% and 69 to >100% for serovar. Enteritidis and L. monocytogenes, respectively, in the cheddar cheese samples. There were no significant differences in recovery efficiency at different inocula levels, and losses to discarded supernatants were always <5%, regardless of dairy product or pathogen. Conclusions:, When followed by pathogen detection using PCR and confirmation by amplicon hybridization, detection limits of 103 and 101 CFU per 11-g sample were achieved for L. monocytogenes and serovar. Enteritidis, respectively, in both product types and without prior cultural enrichment. Significance and Impact of the Study:, This study represents progress toward the rapid and efficient direct detection of pathogens from complex food matrices at detection limits approaching those that might be anticipated in naturally contaminated products. [source] Multiple displacement amplification as a pre-polymerase chain reaction (pre-PCR) to process difficult to amplify samples and low copy number sequences from natural environmentsENVIRONMENTAL MICROBIOLOGY, Issue 7 2005Juan M. Gonzalez Summary Microbial assessment of natural biodiversity is usually achieved through polymerase chain reaction (PCR) amplification. Deoxyribonucleic acid (DNA) sequences from natural samples are often difficult to amplify because of the presence of PCR inhibitors or to the low number of copies of specific sequences. In this study, we propose a non-specific preamplification procedure to overcome the presence of inhibitors and to increase the number of copies prior to carrying out standard amplification by PCR. The pre-PCR step is carried out through a multiple displacement amplification (MDA) technique using random hexamers as priming oligonucleotides and ,29 DNA polymerase in an isothermal, whole-genome amplification reaction. Polymerase chain reaction amplification using specific priming oligonucleotides allows the selection of the sequences of interest after a preamplification reaction from complex environmental samples. The procedure (MDA-PCR) has been tested on a natural microbial community from a hypogean environment and laboratory assemblages of known bacterial species, in both cases targeting the small subunit ribosomal RNA gene sequences. Results from the natural community showed successful amplifications using the two steps protocol proposed in this study while standard, direct PCR amplification resulted in no amplification product. Amplifications from a laboratory assemblage by the two-step proposed protocol were successful at bacterial concentrations ,,10-fold lower than standard PCR. Amplifications carried out in the presence of different concentrations of fulvic acids (a soil humic fraction) by the MDA-PCR protocol generated PCR products at concentrations of fulvic acids over 10-fold higher than standard PCR amplifications. The proposed procedure (MDA-PCR) opens the possibility of detecting sequences represented at very low copy numbers, to work with minute samples, as well as to reduce the negative effects on PCR amplifications of some inhibitory substances commonly found in environmental samples. [source] Construction and Characterization of Porous SiO2/Hydrogel Hybrids as Optical Biosensors for Rapid Detection of BacteriaADVANCED FUNCTIONAL MATERIALS, Issue 14 2010Naama Massad-Ivanir Abstract The use of a new class of hybrid nanomaterials as label-free optical biosensors for bacteria detection (E. coli K12 as a model system) is demonstrated. The hybrids combine a porous SiO2 (PSiO2) optical nanostructure (a Fabry,Pérot thin film) used as the optical transducer element and a hydrogel. The hydrogel, polyacrylamide, is synthesized in situ within the nanostructure inorganic host and conjugated with specific monoclonal antibodies (IgGs) to provide the active component of the biosensor. The immobilization of the IgGs onto the hydrogel via a biotin-streptavidin system is confirmed by fluorescent labeling experiments and reflective interferometric Fourier transform spectroscopy (RIFTS). Additionally, the immobilized IgGs maintain their immunoactivity and specificity when attached to the sensor surface. Exposure of these modified-hybrids to the target bacteria results in "direct cell capture" onto the biosensor surface. These specific binding events induce predictable changes in the thin-film optical interference spectrum of the hybrid. Preliminary studies demonstrate the applicability of these biosensors for the detection of low bacterial concentrations in the range of 103,105 cell mL,1 within minutes. [source] Is Helicobacter pylori a True Microaerophile?HELICOBACTER, Issue 4 2006Stephanie Bury-Moné Abstract Background:, There is no general consensus about the specific oxygen and carbon dioxide requirements of the human pathogen Helicobacter pylori. This bacterium is considered a microaerophile and consequently, it is grown under atmospheres at oxygen tensions 5,19% and carbon dioxide tensions 5,10%, both for clinical and basic and applied research purposes. The current study compared the growth of H. pylori in vitro, under various gas atmospheres, and determined some specific changes in the physiology of bacteria grown under different oxygen partial pressures. Methods:, Measurements of bacterial growth under various conditions were carried out employing classical solid and liquid culture techniques. Enzymatic activities were measured using spectrophotometric assays. Results:,H. pylori and all the other Helicobacter spp. tested had an absolute requirement for elevated carbon dioxide concentrations in the growth atmosphere. In contrast with other Helicobacter spp., H. pylori can tolerate elevated oxygen tensions when grown at high bacterial concentrations. Under 5% CO2, the bacterium showed similar growth in liquid cultures under oxygen tensions from microaerobic (< 5%) to fully aerobic (21%) at cell densities higher than 5 × 105 cfu/ml for media supplemented with horse serum and 5 × 107 cfu/ml for media supplemented with ,-cyclodextrin. Evidence that changes occurred in the physiology of H. pylori was obtained by comparing the activities of ferredoxin:NADH (nicotinamide adenine dinucleotide) oxidoreductases of bacteria grown under microaerobic and aerobic atmospheres. Conclusions:,H. pylori is a capnophile able to grow equally well in vitro under microaerobic or aerobic conditions at high bacterial concentrations, and behaved like oxygen-sensitive microaerophiles at low cell densities. Some characteristics of H. pylori cells grown in vitro under microaerobic conditions appeared to mimic better the physiology of organisms grown in their natural niche in the human stomach. [source] Evaluation of the antimicrobial activity of cationic polyethylenimines on dry surfacesBIOTECHNOLOGY PROGRESS, Issue 5 2008Stephen A. Koplin Abstract The antimicrobial activity of cationic N -dodecyl- N -methylpolyethylenimine (PEI+) against S. aureus, A. baumannii, and E. coli was evaluated when the polymer was applied as a coating to various surfaces using a paint-like procedure. Antimicrobial activity of PEI+ as a function of time was determined using an assay for long-term survival involving placement of single drops of various bacterial concentrations on dry surfaces. These data were compared with an assay method where bacteria were applied by spraying and surfaces were incubated overnight under agar. PEI+-coated surfaces were found to be highly bactericidal after 30 min when bacteria were sprayed onto surfaces. However, when bacteria were applied as single drops, PEI+-coated surfaces were less biocidal at short contact times particularly for A. baumannii and E. coli. The observations are explained in the context of the difference in drying time between drops deposited on uncoated surfaces and PEI+-coated surfaces and the sensitivity of bacterial survival to dehydration. These results demonstrate that PEI+-coated surfaces are not effectively biocidal for some types of bacteria under certain conditions and that the method of assaying bactericidal efficiency can greatly affect the results obtained. [source] | ||||||||||