The Experts below are selected from a list of 33069 Experts worldwide ranked by ideXlab platform
Osami Yagi - One of the best experts on this subject based on the ideXlab platform.
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development of a Most Probable Number method for enumerating denitrifying bacteria by using 96 well microtiter plates and an anaerobic culture system
2003Co-Authors: Satoshi Saitoh, Kazuhiro Iwasaki, Osami YagiAbstract:A new Most-Probable-Number (MPN) method using 96-well microtiter plates was developed to enumerate denitrifying bacteria. In this method, dilutions of samples are added to microtiter plates with a medium and incubated anaerobically using the AnaeroPouch culture system (Mitsubishi Gas Chemical, Inc., Tokyo, Japan). The microtiter plate MPN method gave precise estimates of the population densities of a culture of Paracoccus denitrificans, similar to the estimates obtained using a test tube MPN method and a colony count method. The population densities of denitrifying bacteria in soil samples estimated by the microtiter plate MPN method were higher than the population densities estimated by the test tube MPN method. Because the new method requires less equipment, labor, and time than the conventional test tube method. It will be valuable for estimating the biomass of denitrifying bacteria in natural samples.
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development of a new Most Probable Number method for enumerating methanotrophs using 48 well microtiter plates
2002Co-Authors: Satoshi Saitoh, Kazuhiro Iwasaki, Osami YagiAbstract:We developed a new Most-Probable-Number (MPN) method for enumeration of methanotrophs, using 48-well microtiter plates. With this method, a microtiter plate which has been inoculated with serial dilutions of a sample is placed in a plastic desiccator containing methane and shaken. The proposed method gave estimates of population densities of a pure culture of Methylocystis sp. strain M and of methanotrophs in a paddy soil sample similar to those obtained by a conventional MPN method using serum bottles. The new microtiter plate MPN method with an additional replica-plating technique made it possible to enumerate small populations of methanotrophs in cornfield and forest soils. It also requires less equipment, labor, and time than the conventional serum bottle MPN method.
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Quantitative and specific detection of a trichloroethylene-degrading methanotroph, Methylocystis sp. strain M, by a Most Probable Number-polymerase chain reaction method.
2001Co-Authors: Takeshi Kikuchi, Kazuhiro Iwasaki, Hirofumi Nishihara, Yoshichika Takamura, Osami YagiAbstract:We developed a rapid and specific enumeration method for a trichloroethylene-degrading methanotroph, Methylocystis sp. strain M, based on a Most Probable Number-polymerase chain reaction method for monitoring the bacterium at bioremediation sites. The primers designed for the mmoC gene of the soluble methane monooxygenase gene cluster were specific to strain M. Recovery of the cells with a membrane filter enabled us to detect strain M in trichloroethylene-contaminated groundwater. We used the enumeration method to monitor the Number of strain M cells in effluent from soil columns supplied with trichloroethylene-contaminated groundwater. The Number of strain M cells in the effluent depended on the amount of the strain M inoculated and the Number of cells measured by the Most Probable Number-polymerase chain reaction method was correlated with that measured by a culture method. The detection limit for strain M in effluent detected by MPN-PCR method was 4 to 8×102 cells/ml.
J T Trevors - One of the best experts on this subject based on the ideXlab platform.
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comparison of Most Probable Number pcr and Most Probable Number foci detection method for quantifying infectious cryptosporidium parvum oocysts in environmental samples
2006Co-Authors: Christine M Carey, Hung Lee, J T TrevorsAbstract:Microbial contamination of public water supplies is of significant concern, as numerous outbreaks, including Cryptosporidium, have been reported worldwide. Detection and enumeration of Cryptosporidium parvum oocysts in water supplies is important for the prevention of future cryptosporidiosis outbreaks. In addition to not identifying the oocyst species, the U.S. EPA Method 1622 does not provide information on oocyst viability or infectivity. As such, current detection strategies have been coupled with in vitro culture methods to assess oocyst infectivity. In this study, a Most Probable Number (MPN) method was coupled with PCR (MPN-PCR) to quantify the Number of infectious oocysts recovered from seeded raw water concentrates. The frequency of positive MPN-PCR results decreased as the oocyst Numbers decreased. Similar results were observed when MPN was coupled to the foci detection method (MPN-FDM), which was done for comparison. For both methods, infectious oocysts were not detected below 10(3) seeded oocysts and the MPN-PCR and MPN-FDM estimates for each seed dose were generally within one-log unit of directly enumerated foci of infection. MPN-PCR estimates were 0.25, 0.54, 0 and 0.66 log(10) units higher than MPN-FDM estimates for the positive control, 10(5), 10(4) and 10(3) seed doses, respectively. The results show the MPN-PCR was the better method for the detection of infectious C. parvum oocysts in environmental water samples.
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bioluminescent Most Probable Number method to enumerate lux marked pseudomonas aeruginosa ug2lr in soil
1994Co-Authors: C A Flemming, Hung Lee, J T TrevorsAbstract:Abstract Bioluminescence measured with a luminometer and charge-coupled device was an effective marker in Most-Probable-Number assays for luxAB-marked Pseudomonas aeruginosa UG2Lr in soil. Most-Probable-Number assays with microtiter plate wells and luminometer tubes gave estimates for UG2Lr that were similar to viable colony counts. Both methods detected five cells per g of soil.
Albert D Venosa - One of the best experts on this subject based on the ideXlab platform.
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selective enumeration of aromatic and aliphatic hydrocarbon degrading bacteria by a Most Probable Number procedure
1996Co-Authors: Brian A Wrenn, Albert D VenosaAbstract:A Most-Probable-Number (MPN) procedure was developed to separately enumerate aliphatic and aromatic hydrocarbon degrading bacteria, because Most of the currently available methods are unable to dis...
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selective enumeration of aromatic and aliphatic hydrocarbon degrading bacteria by a Most Probable Number procedure
1996Co-Authors: Brian A Wrenn, Albert D VenosaAbstract:A Most-Probable-Number (MPN) procedure was developed to separately enumerate aliphatic and aromatic hydrocarbon degrading bacteria, because Most of the currently available methods are unable to distinguish between these two groups. Separate 96-well microtiter plates are used to estimate the sizes of these two populations. The alkane-degrader MPN method uses hexadecane as the selective growth substrate and positive wells are detected by reduction of iodonitrotetrazolium violet, which is added after incubation for 2 weeks at 20 °C. Polycyclic aromatic hydrocarbon degraders are grown on a mixture of phenanthrene, anthracene, fluorene, and dibenzothiophene in a second plate. Positive wells turn yellow to greenish-brown from accumulation of the partial oxidation products of the aromatic substrates and they can be scored after a 3-week incubation period. These MPN procedures are accurate and selective. For pure cultures, heterotrophic plate counts on a nonselective medium and the appropriate MPN procedure provi...
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measurement of hydrocarbon degrading microbial populations by a 96 well plate Most Probable Number procedure
1996Co-Authors: John R Haines, Brian A Wrenn, Edith L Holder, Kevin L Strohmeier, R T Herrington, Albert D VenosaAbstract:A 96-well microtiter plate Most-Probable-Number (MPN) procedure was developed to enumerate hydrocarbon-degrading microorganisms. The performance of this method, which uses Number 2 fuel oil (F2) as the selective growth substrate and reduction of iodonitrotetrazolium violet (INT) to detect positive wells, was evaluated by comparison with an established 24-well microtiter plate MPN procedure (the Sheen Screen), which uses weathered North Slope crude oil as the selective substrate and detects positive wells by emulsification or dispersion of the oil. Both procedures gave similar estimates of the hydrocarbon-degrader population densities in several oil-degrading enrichment cultures and sand samples from a variety of coastal sites. Although several oils were effective substrates for the 96-well procedure, the combination of F2 with INT was best, because the color change associated with INT reduction was more easily detected in the small wells than was disruption of the crude oil slick. The method's accuracy was evaluated by comparing hydrocarbon-degrader MPNs with heterotrophic plate counts for several pure and mixed cultures. For some organisms, it seems likely that a single cell cannot initiate sufficient growth to produce a positive result. Thus, this and other hydrocarbon-degrader MPN procedures might underestimate the hydrocarbon-degrading population, even for culturable organisms.
D S Fisher - One of the best experts on this subject based on the ideXlab platform.
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Most Probable Number methodology for quantifying dilute concentrations and fluxes of escherichia coli o157 h7 in surface waters
2009Co-Authors: Michael B Jenkins, Dinku M Endale, D S FisherAbstract:Aims: To better understand the transport and enumeration of dilute densities of Escherichia coli O157:H7 in agricultural watersheds, we developed a culturebased, five tube-multiple dilution Most Probable Number (MPN) method. Methods and Results: The MPN method combined a filtration technique for large volumes of surface water with standard selective media, biochemical and immunological tests, and a TaqMan confirmation step. This method determined E. coli O157:H7 concentrations as low as 0AE1 MPN per litre, with a 95% confidence level of 0AE01–0AE7 MPN per litre. Escherichia coli O157:H7 densities ranged from not detectable to 9 MPN per litre for pond inflow, from not detectable to 0AE9 MPN per litre for pond outflow and from not detectable to 8AE3 MPN per litre for within pond. The MPN methodology was extended to mass flux determinations. Fluxes of E. coli O157:H7 ranged from 10 4 MPN per hour. Conclusion: This culture-based method can detect small Numbers of viable ⁄ culturable E. coli O157:H7 in surface waters of watersheds containing animal agriculture and wildlife. Significance and Impact of the Study: This MPN method will improve our understanding of the transport and fate of E. coli O157:H7 in agricultural watersheds, and can be the basis of collections of environmental E. coli
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Most Probable Number methodology for quantifying dilute concentrations and fluxes of salmonella in surface waters
2008Co-Authors: Michael B Jenkins, Dinku M Endale, D S FisherAbstract:Aims: To better understand and manage the fate and transport of Salmonella in agricultural watersheds, we developed a culture-based, five tube‐four dilution Most Probable Number (MPN) method for enumerating dilute densities of Salmonella in environmental waters. Methods and Results: The MPN method was a combination of a filtration technique for large sample volumes of environmental water, standard selective media for Salmonella and a TaqMan confirmation step. This method has determined the density of Salmonella in 20-l samples of pond inflow and outflow streams as low as 0AE1 MPN l )1 and a low 95% confidence level 0AE015 MPN l )1 . Salmonella densities ranged from not detectable to 0AE55 MPN l )1 for pond inflow samples and from not detectable to 3AE4 MPN l )1 for pond outflow samples. Salmonella densities of pond inflow samples were associated with densities of Escherichia coli and faecal enterococci that indicated stream contamination with faeces and with nondetectable pond outflow densities of the faecal indicator bacteria. The MPN methodology was extended to flux determinations by integrating with volumetric measurements of pond inflow (mean flux of 2AE 5ls )1 ) and outflow (mean flux of 5AE 6ls )1 ). Fluxes of Salmonella ranged from 100 to greater than 10 4 MPN h )1 . Conclusions: This is a culture-based method that can detect small Numbers of Salmonella in environmental waters of watersheds containing animal husbandry and wildlife. Significance and Impact of the Study: Applying this method to environmental waters will improve our understanding of the transport and fate of Salmonella in agricultural watersheds, and can be the basis of valuable collections of environmental Salmonella.
Ellen Raber - One of the best experts on this subject based on the ideXlab platform.
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Most Probable Number rapid viability pcr method to detect viable spores of bacillus anthracis in swab samples
2010Co-Authors: Sonia E Letant, Staci R Kane, Gloria A Murphy, Teneile M Alfaro, Lisa Hodges, Laura J Rose, Ellen RaberAbstract:A comparison of Most-Probable-Number Rapid Viability (MPN RV) PCR and traditional culture methods for the quantification of Bacillus anthracis Sterne spores in macrofoam swabs from a multi-center validation study was performed. The purpose of the study was to compare environmental swab processing methods for recovery, detection, and quantification of viable B. anthracis spores from surfaces. Results show that spore Numbers provided by the MPN RV-PCR method were typically within 1-log of the values from a plate count method for all three levels of spores tested (3.1x10(4), 400, and 40 spores sampled from surfaces with swabs) even in the presence of debris. The MPN method tended to overestimate the expected result, especially at lower spore levels. Blind negative samples were correctly identified using both methods showing a lack of cross contamination. In addition to detecting low levels of spores in environmental conditions, the MPN RV-PCR method is specific, and compatible with automated high-throughput sample processing and analysis protocols, enhancing its utility for characterization and clearance following a biothreat agent release.
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Most Probable Number rapid viability pcr method to detect viable spores of bacillus anthracis in swab samples
2008Co-Authors: Sonia E Letant, Staci R Kane, Gloria A Murphy, Teneile M Alfaro, Lisa Hodges, Laura J Rose, Ellen RaberAbstract:This note presents a comparison of Most-Probable-Number Rapid Viability (MPN-RV) PCR and traditional culture methods for the quantification of Bacillus anthracis Sterne spores in macrofoam swabs generated by the Centers for Disease Control and Prevention (CDC) for a multi-center validation study aimed at testing environmental swab processing methods for recovery, detection, and quantification of viable B. anthracis spores from surfaces. Results show that spore Numbers provided by the MPN RV-PCR method were in statistical agreement with the CDC conventional culture method for all three levels of spores tested (10{sup 4}, 10{sup 2}, and 10 spores) even in the presence of dirt. In addition to detecting low levels of spores in environmental conditions, the MPN RV-PCR method is specific, and compatible with automated high-throughput sample processing and analysis protocols.