Reduced Detectability of Listeria monocytogenes in the Presence of Listeria innocua

2011 ◽  
Vol 74 (8) ◽  
pp. 1282-1287 ◽  
Author(s):  
ULRIKE ZITZ ◽  
MARIJA ZUNABOVIC ◽  
KONRAD J. DOMIG ◽  
PETER-THEODOR WILRICH ◽  
WOLFGANG KNEIFEL
Keyword(s):  
Listeria Monocytogenes ◽  
Food Safety ◽  
Limit Of Detection ◽  
False Negative ◽  
Test Methods ◽  
Probability Of Detection ◽  
Listeria Innocua ◽  
Selective Enrichment ◽  
Detection Techniques ◽  
Automated Method

Recent foodborne crises have demonstrated the importance of monitoring food safety. In terms of microbiological criteria, food safety requires the reliable detection of pathogens such as Listeria monocytogenes along the food chain by appropriate analytical methods. However, indications exist that accompanying Listeria innocua strains suppress the growth of L. monocytogenes during selective enrichment, which may cause reduced or even inhibited detection. To study these effects, the limit of detection of L. monocytogenes was investigated in the presence of L. innocua using the International Organization for Standardization standard method ISO 11290-1 and the VIDAS LDUO system, an automated method based on enzyme-linked fluorescence technology. The challenge was to provide low initial Listeria concentrations at sufficient precision to quantify the influence on the probability of detection of L. monocytogenes. The application of reference materials appropriate for quantitative test methods and a standardized dilution procedure were necessary to ensure accurate CFU levels of defined proportions of mixtures of both Listeria species. During selective enrichment, overgrowth of L. monocytogenes by L. innocua could be confirmed, leading to high rates of false-negative results. Moreover, with both methods, a significant decrease in the detectability of L. monocytogenes could be quantified at ratios of 2:1 at very low concentrations representative of natural contamination levels often found in foods and environments. It is concluded that there is a need to improve existing procedures with respect to selective enrichment, as well as the detection techniques.

scholarly journals Persistence of Listeria innocua on Fresh Apples During Long-Term Controlled Atmosphere Cold Storage with Postharvest Fungal Decay

2021 ◽  
Author(s):  
Alexis Hamilton ◽  
Blanca Ruiz-Llacsahuanga ◽  
Manoella Mendoza ◽  
James Mattheis ◽  
Ines Hanrahan ◽  
...  
Keyword(s):  
Food Safety ◽  
Cold Storage ◽  
Limit Of Detection ◽  
Growth Patterns ◽  
Listeria Innocua ◽  
Penicillium Expansum ◽  
Controlled Atmosphere ◽  
Storage Duration ◽  
Apple Variety

Recent apple-related recall and outbreak events have exposed a need for better food safety controls along the supply chain. Following harvest apples can be stored under a controlled atmosphere for up to one year after harvest before packing and distribution, making the crop susceptible to many opportunities for contamination that increase the quantity of postharvest losses. Botrytis cinerea (BC) and Penicillium expansum (PE) cause significant rot-associated losses to the apple industry. These fungi can colonize and destroy apple tissue as storage duration increases, which may also impact the growth of saprophytic foodborne pathogens like Listeria monocytogenes . Thus the objective of this study was to observe population changes of Listeria innocua (LI) as a surrogate for L. monocytogenes on apples inoculated with BC or PE under long-term controlled atmosphere cold storage conditions to identify the effect of postharvest mold growth on growth patterns of a food safety-relevant microorganism. ‘Gala’ and ‘WA 38’ apples (n = 1,080) were harvested, treated with pyrimethanil, and inoculated with LI only, or LI and one of the mold species on wounded and unwounded portions of the apple equator. Apples were treated with 1-methylcyclopropene and stored at a controlled atmosphere (2kPa O 2 , 1kPa CO 2 , 1°C) for 1 week and 1, 3, 6, 9 and 11 months before enumeration. After three months LI consistently fell below the limit of detection (2.35 log CFU/g) and samples were enriched following a modified BAM method with PCR confirmation. Listeria persistence was dependent on the storage duration and type of fungal contamination ( p < 0.05). Surface wounding may impact these trends, depending on the apple variety.   Prevalence of LI was greater in ‘Gala’ apples. Future studies should more closely examine the interactions on the fruit surface that occur during the seemingly critical timeframe of three-to-six months in storage.

Flow Cytometry for Automated Analysis of Milk Containing Listeria monocytogenes

1988 ◽  
Vol 71 (3) ◽  
pp. 655-658
Author(s):  
Catherine W Donnelly ◽  
Gregory J Baigent ◽  
Elizabeth H Briggs
Keyword(s):  
Flow Cytometry ◽  
Listeria Monocytogenes ◽  
False Positive Rate ◽  
False Negative ◽  
False Negative Rate ◽  
Automated Analysis ◽  
Raw Milk ◽  
Fluorescence Labeling ◽  
Selective Enrichment ◽  
Milk Samples

Abstract This paper describes the application of flow cytometry to the analysis of milk for the presence of Listeria monocytogenes. A total of 939 raw milk samples from 54 farms were analyzed for the presence of L. monocytogenes by cold enrichment vs flow cytometry, which incorporated a selective enrichment step. Analyzed samples consisted of milk from individual farm bulk tanks; string samples which represented milk from 25 to 40 cows; and combination samples representing milk from 200 cows. Raw milk samples were directly enriched for 24 h at 37°C and analyzed by flow cytometry following fluorescence labeling of bacterial populations, or were cold enriched for 1 month at 4°C. Following cold enrichment, samples were streaked to McBride's Listeria agar, and suspect colonies were biochemically identified as L. monocytogenes. L. monocytogenes was isolated from only 15 of the analyzed samples, all of which were common to one farm. Results of flow cytometry analysis yielded a 5.86% false positive rate and a 0.53% false negative rate when compared with culture procedures

scholarly journals Direct and rapid measurement of hydrogen peroxide in human blood using a microfluidic device

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
R. Gaikwad ◽  
P. R. Thangaraj ◽  
A. K. Sen
Keyword(s):  
Hydrogen Peroxide ◽  
Microfluidic Device ◽  
Human Blood ◽  
Blood Cells ◽  
Limit Of Detection ◽  
Point Of Care ◽  
Time Interval ◽  
Fluorescence Measurement ◽  
Rapid Measurement ◽  
Automated Method

AbstractThe levels of hydrogen peroxide ($${\mathrm{H}}_{2}{\mathrm{O}}_{2}$$ H 2 O 2 ) in human blood is of great relevance as it has emerged as an important signalling molecule in a variety of disease states. Fast and reliable measurement of $${\mathrm{H}}_{2}{\mathrm{O}}_{2}$$ H 2 O 2 levels in the blood, however, continues to remain a challenge. Herein we report an automated method employing a microfluidic device for direct and rapid measurement of $${\mathrm{H}}_{2}{\mathrm{O}}_{2}$$ H 2 O 2 in human blood based on laser-induced fluorescence measurement. Our study delineates the critical factors that affect measurement accuracy—we found blood cells and soluble proteins significantly alter the native $${\mathrm{H}}_{2}{\mathrm{O}}_{2}$$ H 2 O 2 levels in the time interval between sample withdrawal and detection. We show that separation of blood cells and subsequent dilution of the plasma with a buffer at a ratio of 1:6 inhibits the above effect, leading to reliable measurements. We demonstrate rapid measurement of $${\mathrm{H}}_{2}{\mathrm{O}}_{2}$$ H 2 O 2 in plasma in the concentration range of 0–49 µM, offering a limit of detection of 0.05 µM, a sensitivity of 0.60 µM−1, and detection time of 15 min; the device is amenable to the real-time measurement of $${\mathrm{H}}_{2}{\mathrm{O}}_{2}$$ H 2 O 2 in the patient’s blood. Using the linear correlation obtained with known quantities of $${\mathrm{H}}_{2}{\mathrm{O}}_{2}$$ H 2 O 2 , the endogenous $${\mathrm{H}}_{2}{\mathrm{O}}_{2}$$ H 2 O 2 concentration in the blood of healthy individuals is found to be in the range of 0.8–6 µM. The availability of this device at the point of care will have relevance in understanding the role of $${\mathrm{H}}_{2}{\mathrm{O}}_{2}$$ H 2 O 2 in health and disease.

scholarly journals Digital Droplet PCR for SARS-CoV-2 Resolves Borderline Cases

2021 ◽  
Author(s):  
Jing Xu ◽  
Timothy Kirtek ◽  
Yan Xu ◽  
Hui Zheng ◽  
Huiyu Yao ◽  
...  
Keyword(s):  
Polymerase Chain Reaction ◽  
Limit Of Detection ◽  
False Negative ◽  
Accurate Method ◽  
Viral Envelope ◽  
Chain Reaction ◽  
Borderline Cases ◽  
Droplet Pcr ◽  
Polymerase Chain ◽  
Digital Polymerase Chain Reaction

Abstract Objectives The Bio-Rad SARS-CoV-2 ddPCR Kit (Bio-Rad Laboratories) was the first droplet digital polymerase chain reaction (ddPCR) assay to receive Food and Drug Administration (FDA) Emergency Use Authorization approval, but it has not been evaluated clinically. We describe the performance of ddPCR—in particular, its ability to confirm weak-positive severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) results. Methods We clinically validated the Bio-Rad Triplex Probe ddPCR Assay. The limit of detection was determined by using serial dilutions of SARS-CoV-2 RNA in an artificial viral envelope. The ddPCR assay was performed according to the manufacturer’s specifications on specimens confirmed to be positive (n = 48) or negative (n = 30) by an FDA-validated reverse transcription–polymerase chain reaction assay on the m2000 RealTime system (Abbott). Ten borderline positive cases were also evaluated. Results The limit of detection was 50 copies/mL (19 of 20 positive). Forty-seven specimens spanning a range of quantification cycles (2.9-25.9 cycle numbers) were positive by this assay (47 of 48; 97.9% positive precent agreement), and 30 negative samples were confirmed as negative (30 of 30; 100% negative percent agreement). Nine of 10 borderline cases were positive when tested in triplicate. Conclusions The ddPCR of SARS-CoV-2 is an accurate method, with superior sensitivity for viral RNA detection. It could provide definitive evaluation of borderline positive cases or suspected false-negative cases.

Automation and validation of a MALDI-TOF MS (Mass-Fix) replacement of immunofixation electrophoresis in the clinical lab

2021 ◽  
Vol 59 (1) ◽  
pp. 155-163
Author(s):  
Mindy Kohlhagen ◽  
Surendra Dasari ◽  
Maria Willrich ◽  
MeLea Hetrick ◽  
Brian Netzel ◽  
...  
Keyword(s):  
Limit Of Detection ◽  
Turnaround Time ◽  
Automated System ◽  
Serum Samples ◽  
Maldi Tof Ms ◽  
Maldi Tof ◽  
Automated Method ◽  
Positive Specimen ◽  
Specimen Identification ◽  
Tof Ms

AbstractObjectivesA matrix assisted laser desorption ionization time of flight mass spectrometry (MALDI-TOF MS) method (Mass-Fix) as a replacement for gel-based immunofixation (IFE) has been recently described. To utilize Mass-Fix clinically, a validated automated method was required. Our aim was to automate the pre-analytical processing, improve positive specimen identification and ergonomics, reduce paper data storage and increase resource utilization without increasing turnaround time.MethodsSerum samples were batched and loaded onto a liquid handler along with reagents and a barcoded sample plate. The pre-analytical steps included: (1) Plating immunopurification beads. (2) Adding 10 μl of serum. (3) Bead washing. (4) Eluting the immunoglobulins (Igs), and reducing to separate the heavy and light Ig chains. The resulting plate was transferred to a second low-volume liquid handler for MALDI plate spotting. MALDI-TOF mass spectra were collected. Integrated in-house developed software was utilized for sample tracking, driving data acquisition, data analysis, history tracking, and result reporting. A total of 1,029 residual serum samples were run using the automated system and results were compared to prior electrophoretic results.ResultsThe automated Mass-Fix method was capable of meeting the validation requirements of concordance with IFE, limit of detection (LOD), sample stability and reproducibility with a low repeat rate. Automation and integrated software allowed a single user to process 320 samples in an 8 h shift. Software display facilitated identification of monoclonal proteins. Additionally, the process maintains positive specimen identification, reduces manual pipetting, allows for paper free tracking, and does not significantly impact turnaround time (TAT).ConclusionsMass-Fix is ready for implementation in a high-throughput clinical laboratory.

scholarly journals High sensitivity sanger sequencing detection of BRAF mutations in metastatic melanoma FFPE tissue specimens

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Lauren Y. Cheng ◽  
Lauren E. Haydu ◽  
Ping Song ◽  
Jianyi Nie ◽  
Michael T. Tetzlaff ◽  
...  
Keyword(s):  
Sanger Sequencing ◽  
Limit Of Detection ◽  
False Negative ◽  
High Sensitivity ◽  
Ffpe Tissue ◽  
Braf Mutations ◽  
Tissue Samples ◽  
Ffpe Samples ◽  
Droplet Pcr ◽  
Tissue Specimens

AbstractMutations in the BRAF gene at or near the p. V600 locus are informative for therapy selection, but current methods for analyzing FFPE tissue DNA generally have a limit of detection of 5% variant allele frequency (VAF), or are limited to the single variant (V600E). These can result in false negatives for samples with low VAFs due to low tumor content or subclonal heterogeneity, or harbor non-V600 mutations. Here, we show that Sanger sequencing using the NuProbe VarTrace BRAF assay, based on the Blocker Displacement Amplification (BDA) technology, is capable of detecting BRAF V600 mutations down to 0.20% VAF from FFPE lymph node tissue samples. Comparison experiments on adjacent tissue sections using BDA Sanger, immunohistochemistry (IHC), digital droplet PCR (ddPCR), and NGS showed 100% concordance among all 4 methods for samples with BRAF mutations at ≥ 1% VAF, though ddPCR did not distinguish the V600K mutation from the V600E mutation. BDA Sanger, ddPCR, and NGS (with orthogonal confirmation) were also pairwise concordant for lower VAF mutations down to 0.26% VAF, but IHC produced a false negative. Thus, we have shown that Sanger sequencing can be effective for rapid detection and quantitation of multiple low VAF BRAF mutations from FFPE samples. BDA Sanger method also enabled detection and quantitation of less frequent, potentially actionable non-V600 mutations as demonstrated by synthetic samples.

Changes in Aerobic Plate and Escherichia coli–Coliform Counts and in Populations of Inoculated Foodborne Pathogens on Inshell Walnuts during Storage

2016 ◽  
Vol 79 (7) ◽  
pp. 1143-1153 ◽  
Author(s):  
JOHN C. FRELKA ◽  
GORDON R. DAVIDSON ◽  
LINDA J. HARRIS
Keyword(s):  
Escherichia Coli ◽  
Listeria Monocytogenes ◽  
Relative Humidity ◽  
Low Temperature ◽  
Foodborne Pathogens ◽  
Limit Of Detection ◽  
Sampling Time ◽  
E Coli ◽  
Plate Counts ◽  
Forced Air

ABSTRACT After harvest, inshell walnuts are dried using low-temperature forced air and are then stored in bins or silos for up to 1 year. To better understand the survival of bacteria on inshell walnuts, aerobic plate counts (APCs) and Escherichia coli–coliform counts (ECCs) were evaluated during commercial storage (10 to 12°C and 63 to 65% relative humidity) over 9 months. APCs decreased by 1.4 to 2.0 log CFU per nut during the first 5 months of storage, and ECCs decreased by 1.3 to 2.2 log CFU per nut in the first month of storage. Through the remaining 4 to 8 months of storage, APCs and ECCs remained unchanged (P > 0.05) or decreased by <0.15 log CFU per nut per month. Similar trends were observed on kernels extracted from the inshell walnuts. APCs and ECCs were consistently and often significantly higher on kernels extracted from visibly broken inshell walnuts than on kernels extracted from visibly intact inshell walnuts. Parameters measured in this study were used to determine the survival of five-strain cocktails of E. coli O157:H7, Listeria monocytogenes, and Salmonella inoculated onto freshly hulled inshell walnuts (~8 log CFU/g) after simulated commercial drying (10 to 12 h; 40°C) and simulated commercial storage (12 months at 10°C and 65% relative humidity). Populations declined by 2.86, 5.01, and 4.40 log CFU per nut for E. coli O157:H7, L. monocytogenes, and Salmonella, respectively, after drying and during the first 8 days of storage. Salmonella populations changed at a rate of −0.33 log CFU per nut per month between days 8 and 360, to final levels of 2.83 ± 0.79 log CFU per nut. E. coli and L. monocytogenes populations changed by −0.17 log CFU per nut per month and −0.26 log CFU per nut per month between days 8 and 360, respectively. For some samples, E. coli or L. monocytogenes populations were below the limit of detection by plating (0.60 log CFU per nut) by day 183 or 148, respectively; at least one of the six samples was positive at each subsequent sampling time by either plating or by enrichment.

scholarly journals Thermal inactivation and growth potential of Listeria monocytogenes in smoked tench

2016 ◽  
Vol 5 (3) ◽  
Author(s):  
Raffaella Branciari ◽  
Andrea Valiani ◽  
Raffaella Franceschini ◽  
David Ranucci ◽  
Alessia Lupattelli ◽  
...  
Keyword(s):  
Experimental Study ◽  
Listeria Monocytogenes ◽  
Thermal Inactivation ◽  
Challenge Test ◽  
Listeria Innocua ◽  
Growth Potential ◽  
Post Processing ◽  
Colony Forming Unit ◽  
Post Process ◽  
Hygiene Measures

An experimental study for the evaluation of <em>Listeria monocytogenes</em> inactivation during a hot smoking process in tench was performed using <em>Listeria innocua</em> strains. Furthermore, the survival of <em>L. monocytogenes</em> in smoked tench was determined after post-processing in contaminated samples, evaluating the growth potential during storage. <em>L. innocua</em> was not detected after the smoking process. In the challenge test, the growth potential of <em>L. monocytogenes</em> was 5.68 log colony forming unit g<sup>−1</sup>. The results showed that hot smoking at an inner temperature around 72°C is able to eliminate the microorganism. Nevertheless, the product is able to support the growth of the pathogen if post-process contamination occurs, as the food is suitable for <em>Listeria</em> multiplication. Product recontamination should be prevented by means of appropriate application of hygiene measures.

scholarly journals An overview of Listeria monocytogenes contamination in ready to eat meat, dairy and fishery foods

2017 ◽  
Vol 47 (2) ◽  
Author(s):  
Carla Susana Rodrigues ◽  
Cláudia Valéria Gonçalves Cordeiro de Sá ◽  
Cristiano Barros de Melo
Keyword(s):  
Public Health ◽  
Listeria Monocytogenes ◽  
Sodium Chloride ◽  
Food Safety ◽  
Pregnant Women ◽  
Foodborne Pathogen ◽  
Chloride Content ◽  
The Elderly ◽  
Fishery Products ◽  
Serious Disease

ABSTRACT: Listeria monocytogenes is a relevant foodborne pathogen in public health, responsible for outbreaks of listeriosis often associated to the consumption of ready to eat meat, dairy and fishery products. Listeriosis is a serious disease that can lead to death and mainly affect children, the elderly and immunocompromised individuals. In pregnant women causes abortion or neonatal listeriosis. In Brazil, ready to eat food are appreciated and increasingly consumed by the population. Furthermore, products such as sausages, bologna, hams and cheeses have characteristics such as pH, Aw and sodium chloride content that favor the development of L. monocytogenes during their shelf life. The purpose of this paper was to present an overview of L. monocytogenes contamination in different meat, dairy and fishery products that are ready for consumption and thereby support the adoption of strategies to mitigate this risk, contributing to achieve the appropriate level protection for the consumers and thus strengthen Brazil's food safety system.

Sign in / Sign up

Export Citation Format

Share Document