scholarly journals Rapid Quantification of Viable Campylobacter Bacteria on Chicken Carcasses, Using Real-Time PCR and Propidium Monoazide Treatment, as a Tool for Quantitative Risk Assessment

2010 ◽  
Vol 76 (15) ◽  
pp. 5097-5104 ◽  
Author(s):  
M. H. Josefsen ◽  
C. L�fstr�m ◽  
T. B. Hansen ◽  
L. S. Christensen ◽  
J. E. Olsen ◽  
...  
Keyword(s):  
Risk Assessment ◽  
Real Time ◽  
Control Strategies ◽  
Quantitative Risk Assessment ◽  
Quantitative Detection ◽  
Amplification Efficiency ◽  
Sample Treatment ◽  
Propidium Monoazide ◽  
Real Time Quantitative Pcr ◽  
Chicken Carcass

ABSTRACT A number of intervention strategies against Campylobacter-contaminated poultry focus on postslaughter reduction of the number of cells, emphasizing the need for rapid and reliable quantitative detection of only viable Campylobacter bacteria. We present a new and rapid quantitative approach to the enumeration of food-borne Campylobacter bacteria that combines real-time quantitative PCR (Q-PCR) with simple propidium monoazide (PMA) sample treatment. In less than 3 h, this method generates a signal from only viable and viable but nonculturable (VBNC) Campylobacter bacteria with an intact membrane. The method's performance was evaluated by assessing the contributions to variability by individual chicken carcass rinse matrices, species of Campylobacter, and differences in efficiency of DNA extraction with differing cell inputs. The method was compared with culture-based enumeration on 50 naturally infected chickens. The cell contents correlated with cycle threshold (CT ) values (R 2 = 0.993), with a quantification range of 1 � 102 to 1 � 107 CFU/ml. The correlation between the Campylobacter counts obtained by PMA-PCR and culture on naturally contaminated chickens was high (R 2 = 0.844). The amplification efficiency of the Q-PCR method was not affected by the chicken rinse matrix or by the species of Campylobacter. No Q-PCR signals were obtained from artificially inoculated chicken rinse when PMA sample treatment was applied. In conclusion, this study presents a rapid tool for producing reliable quantitative data on viable Campylobacter bacteria in chicken carcass rinse. The proposed method does not detect DNA from dead Campylobacter bacteria but recognizes the infectious potential of the VBNC state and is thereby able to assess the effect of control strategies and provide trustworthy data for risk assessment.

Quantitative Risk Assessment of Listeriosis-Associated Deaths Due to Listeria monocytogenes Contamination of Deli Meats Originating from Manufacture and Retail

2010 ◽  
Vol 73 (4) ◽  
pp. 620-630 ◽  
Author(s):  
ABANI K. PRADHAN ◽  
RENATA IVANEK ◽  
YRJÖ T. GRÖHN ◽  
ROBERT BUKOWSKI ◽  
IFIGENIA GEORNARAS ◽  
...  
Keyword(s):  
Risk Assessment ◽  
Listeria Monocytogenes ◽  
Relative Risk ◽  
Storage Temperature ◽  
Control Strategies ◽  
Quantitative Risk Assessment ◽  
Sensitivity Analyses ◽  
Effective Control ◽  
Lag Phase ◽  
Reported Data

The objective of this study was to estimate the relative risk of listeriosis-associated deaths attributable to Listeria monocytogenes contamination in ham and turkey formulated without and with growth inhibitors (GIs). Two contamination scenarios were investigated: (i) prepackaged deli meats with contamination originating solely from manufacture at a frequency of 0.4% (based on reported data) and (ii) retail-sliced deli meats with contamination originating solely from retail at a frequency of 2.3% (based on reported data). Using a manufacture-to-consumption risk assessment with product-specific growth kinetic parameters (i.e., lag phase and exponential growth rate), reformulation with GIs was estimated to reduce human listeriosis deaths linked to ham and turkey by 2.8- and 9-fold, respectively, when contamination originated at manufacture and by 1.9- and 2.8-fold, respectively, for products contaminated at retail. Contamination originating at retail was estimated to account for 76 and 63% of listeriosis deaths caused by ham and turkey, respectively, when all products were formulated without GIs and for 83 and 84% of listeriosis deaths caused by ham and turkey, respectively, when all products were formulated with GIs. Sensitivity analyses indicated that storage temperature was the most important factor affecting the estimation of per annum relative risk. Scenario analyses suggested that reducing storage temperature in home refrigerators to consistently below 7°C would greatly reduce the risk of human listeriosis deaths, whereas reducing storage time appeared to be less effective. Overall, our data indicate a critical need for further development and implementation of effective control strategies to reduce L. monocytogenes contamination at the retail level.

Propidium monoazide combined with real-time quantitative PCR underestimates heat-killed Listeria innocua

2011 ◽  
Vol 85 (2) ◽  
pp. 164-169 ◽  
Author(s):  
Trond Løvdal ◽  
Maria Befring Hovda ◽  
Benny Björkblom ◽  
Simon G. Møller
Keyword(s):  
Real Time ◽  
Quantitative Pcr ◽  
Listeria Innocua ◽  
Propidium Monoazide ◽  
Real Time Quantitative Pcr

scholarly journals Implications of Limits of Detection of Various Methods for Bacillus anthracis in Computing Risks to Human Health

2009 ◽  
Vol 75 (19) ◽  
pp. 6331-6339 ◽  
Author(s):  
Amanda B. Herzog ◽  
S. Devin McLennan ◽  
Alok K. Pandey ◽  
Charles P. Gerba ◽  
Charles N. Haas ◽  
...  
Keyword(s):  
Risk Assessment ◽  
Bacillus Anthracis ◽  
Real Time ◽  
Real Time Pcr ◽  
Limit Of Detection ◽  
Quantitative Risk Assessment ◽  
Detection Methods ◽  
Enzyme Linked Immunosorbent Assay ◽  
Environmental Matrices ◽  
Limits Of Detection

ABSTRACT Used for decades for biological warfare, Bacillus anthracis (category A agent) has proven to be highly stable and lethal. Quantitative risk assessment modeling requires descriptive statistics of the limit of detection to assist in defining the exposure. Furthermore, the sensitivities of various detection methods in environmental matrices are vital information for first responders. A literature review of peer-reviewed journal articles related to methods for detection of B. anthracis was undertaken. Articles focused on the development or evaluation of various detection approaches, such as PCR, real-time PCR, immunoassay, etc. Real-time PCR and PCR were the most sensitive methods for the detection of B. anthracis, with median instrument limits of detection of 430 and 440 cells/ml, respectively. There were very few peer-reviewed articles on the detection methods for B. anthracis in the environment. The most sensitive limits of detection for the environmental samples were 0.1 CFU/g for soil using PCR-enzyme-linked immunosorbent assay (ELISA), 17 CFU/liter for air using an ELISA-biochip system, 1 CFU/liter for water using cultivation, and 1 CFU/cm2 for stainless steel fomites using cultivation. An exponential dose-response model for the inhalation of B. anthracis estimates of risk at concentrations equal to the environmental limit of detection determined the probability of death if untreated to be as high as 0.520. Though more data on the environmental limit of detection would improve the assumptions made for the risk assessment, this study's quantification of the risk posed by current limitations in the knowledge of detection methods should be considered when employing those methods in environmental monitoring and cleanup strategies.

scholarly journals Propidium monoazide combined with real-time quantitative PCR to quantify viable Alternaria spp. contamination in tomato products

2013 ◽  
Vol 165 (3) ◽  
pp. 214-220 ◽  
Author(s):  
Ana Crespo-Sempere ◽  
Núria Estiarte ◽  
Sonia Marín ◽  
Vicente Sanchis ◽  
Antonio J. Ramos
Keyword(s):  
Real Time ◽  
Quantitative Pcr ◽  
Propidium Monoazide ◽  
Real Time Quantitative Pcr ◽  
Tomato Products

scholarly journals Rapidly quantitative detection of Nosema ceranae in honeybees using ultra-rapid real-time quantitative PCR

2021 ◽  
Vol 22 ◽  
Author(s):  
A-Tai Truong ◽  
Sedat Sevin ◽  
Seonmi Kim ◽  
Mi-Sun Yoo ◽  
Yun Sang Cho ◽  
...  
Keyword(s):  
Real Time ◽  
Quantitative Pcr ◽  
Nosema Ceranae ◽  
Quantitative Detection ◽  
Real Time Quantitative Pcr

Rapid Detection of DNA Methylation with a Novel Real-Time Fluorescence Recombinase-Aided Amplification Assay

2021 ◽  
Vol 17 (7) ◽  
pp. 1364-1370
Author(s):  
Ziyu He ◽  
Zengrui Tong ◽  
Boyu Tan ◽  
Xuliang He ◽  
Tao Zhang ◽  
...  
Keyword(s):  
Dna Methylation ◽  
Real Time ◽  
Quantitative Detection ◽  
Amplification Efficiency ◽  
Clinical Settings ◽  
Specific Sequence ◽  
Fluorogenic Probe ◽  
Detection Assay ◽  
Amplification Assay ◽  
Entire Experiment

Researchers have conducted in-depth research on DNA methylation mechanism, which is related to various diseases such as deficiency of imprinted gene and occurrence of tumors. This study provides a novel rapid quantitative detection assay and real-time fluorescence recombinase-aided amplification assay (RAA) for DNA methylation. Firstly, specific sequence of methylation genes was chosen and primers and fluorogenic probe for RAA experiment were designed and synthesized. Lastly, these amplification products were proven by sequencing and analysis. Results showed that the amplification efficiency and template concentration of RAA had linear dependent (R2 > 95%) when the concentration range was 4.64×108 copies/μL˜4.64×104 copies/μL. The test assay can also detect positive samples when the template concentration is below 4.64×104 copies/μL. Remarkably, the entire experiment process only takes 15–20 minutes, so it is beneficial for rapid bedside simple screening of some special DNA methylation sites, such as detection of resistance genes. In a word, this method has very great potential for diseases with DNA methylation in clinical settings, especially if methylation analysis needs to be done quickly and easily.

scholarly journals Direct Quantitative Detection and Identification of Lactococcal Bacteriophages from Milk and Whey by Real-Time PCR: Application for the Detection of Lactococcal Bacteriophages in Goat's Raw Milk Whey in France

2011 ◽  
Vol 2011 ◽  
pp. 1-9 ◽  
Author(s):  
Mai Huong Ly-Chatain ◽  
Loïc Durand ◽  
Véronique Rigobello ◽  
Annabelle Vera ◽  
Yann Demarigny
Keyword(s):  
Real Time ◽  
Real Time Pcr ◽  
Direct Detection ◽  
Raw Milk ◽  
Quantitative Detection ◽  
Amplification Efficiency ◽  
The Real ◽  
Milk Whey ◽  
Detection And Identification ◽  
Milk Fermentation

The presence ofLactococcusbacteriophages in milk can partly or completely inhibit milk fermentation. To prevent the problems associated with the bacteriophages, the real-time PCR was developed in this study for direct detection from whey and milk of three main groups ofLactococcusbacteriophages, c2, 936, and P335. The optimization of DNA extraction protocol from complex matrices such as whey and milk was optimized allowed the amplification of PCR without any matrix and nontarget contaminant interference. The real-time PCR program was specific and with the detection limit of 102PFU/mL. The curve slopes were −3.49, −3.69, and −3.45 with the amplification efficiency estimated at 94%, 94%, and 98% and the correlation coefficient () of 0.999, 0.999, and 0.998 for c2, 936 and P335 group, respectively. This method was then used to detect the bacteriophages in whey and goat's raw milk coming from three farms located in the Rhône-Alpes region (France).

scholarly journals A Data-Driven Approach Based on Multivariate Copulas for Quantitative Risk Assessment of Concrete Dam

2019 ◽  
Vol 7 (10) ◽  
pp. 353 ◽  
Author(s):  
Chenfei Shao ◽  
Chongshi Gu ◽  
Zhenzhu Meng ◽  
Yating Hu
Keyword(s):  
Risk Assessment ◽  
Real Time ◽  
Risk Ratio ◽  
Copula Function ◽  
Quantitative Risk Assessment ◽  
Data Driven ◽  
Data Series ◽  
Monitoring Point ◽  
Risk Probability ◽  
Multivariate Copula

Risk assessment of dam’s running status is an important part of dam management. A data-driven method based on monitored displacement data has been applied in risk assessment, owing to its easy operation and real-time analysis. However, previous data-driven methods considered displacement data series at each monitoring point as an independent variable and assessed the running status of each monitoring point separately, without considering the correlation between displacement of different monitoring points. In addition, previous studies assessed the dam’s running status qualitatively, without quantifying the risk probability. To solve the above two issues, a displacement-data driven method based on a multivariate copula function is proposed in this paper. Multivariate copula functions can construct a joint distribution which reveals the relevance structure of random variables. We assumed that the risk probability of each dam section is independent and took monitoring points at one dam section as examples. Starting from the risk assessment of single monitoring points, we calculated the residual between the monitored displacement data and the modelled data estimated by the statistical model, and built a risk ratio function based on the residual. Then, using the multivariate copula function, we obtained a combined risk ratio of multi-monitoring points which took the correlation between each monitoring point into account. Finally, a case study was provided. The proposed method not only quantitatively assessed the probability of the real-time dam risk but also considered the correlation between the displacement data of different monitoring points.

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