Evaluation of Environmental Sampling Methods and Rapid Detection Assays for Recovery and Identification of Listeria spp. from Meat Processing Facilities

2009 ◽  
Vol 72 (4) ◽  
pp. 696-701 ◽  
Author(s):  
JOVANA KOVAČEVIĆ ◽  
VALERIE M. BOHAYCHUK ◽  
PABLO ROMERO BARRIOS ◽  
GARY E. GENSLER ◽  
DEANA L. ROLHEISER ◽  
...  
Keyword(s):  
Sampling Methods ◽  
Culture Method ◽  
Detection Methods ◽  
Lower Sensitivity ◽  
Meat Processing ◽  
Cotton Swab ◽  
Conventional Culture ◽  
Specificity And Sensitivity ◽  
Highly Sensitive ◽  
Labor Inputs

Studies that isolated Listeria spp. from the environment of two meat processing facilities were conducted. Samples were collected in the processing environment of the facilities with three different sampling methods (cotton swab, sterile sponge, and composite-ply tissues) to evaluate their ability to recover Listeria spp. A total of 240 samples for each sampling method were collected and tested. The cotton swab method of sampling was significantly (P < 0.01) less efficient in recovery of Listeria spp. than the sterile-sponge and composite-ply tissue methods, which were similar (P > 0.05) in their ability to recover Listeria spp. The specificity and sensitivity of four detection methods (conventional culture, Petrifilm Environmental Listeria Plates [ELP], lateral-flow immunoprecipitation [LFI], and automated PCR) were evaluated for identification of Listeria spp. Facilities were visited until a minimum of 100 positive and 100 negative samples per detection method were collected. The LFI and PCR methods were highly sensitive (95.5 and 99.1%, respectively) and specific (100%) relative to the culture method. The ELP method was significantly less efficient (P < 0.01) than LFI and PCR in detection of Listeria spp., with lower sensitivity (50.6%) and specificity (91.5%). Kappa values indicated excellent agreement of the LFI and PCR assays and moderate agreement of the ELP method to the culture method. Overall, ELP was easy to use but less efficient in detection of Listeria spp. from environmental samples, while the LFI and PCR methods were found to be excellent alternatives to culture, considering performance and time and labor inputs.

A Real-Time PCR Assay for the Detection of Salmonella in a Wide Variety of Food and Food-Animal Matrices†

2007 ◽  
Vol 70 (5) ◽  
pp. 1080-1087 ◽  
Author(s):  
V. M. BOHAYCHUK ◽  
G. E. GENSLER ◽  
M. E. McFALL ◽  
R. K. KING ◽  
D. G. RENTER
Keyword(s):  
Real Time ◽  
Real Time Pcr ◽  
Culture Method ◽  
Pcr Assay ◽  
Culture Methods ◽  
The Real ◽  
Conventional Culture ◽  
Food Animal ◽  
Highly Sensitive ◽  
Field Samples

Conventional culture methods have traditionally been considered the “gold standards” for the isolation and identification of foodborne pathogens. However, culture methods are labor-intensive and time-consuming. We have developed a real-time PCR assay for the detection of Salmonella in a variety of food and food-animal matrices. The real-time PCR assay incorporates both primers and hybridization probes based on the sequence of the Salmonella invA gene and uses fluorescent resonance energy transfer technology to ensure highly sensitive and specific results. This method correctly classified 51 laboratory isolates of Salmonella and 28 non-Salmonella strains. The method was also validated with a large number of field samples that consisted of porcine feces and cecal contents, pork carcasses, bovine feces and beef carcasses, poultry cecal contents and carcasses, equine feces, animal feeds, and various food products. The samples (3,388) were preenriched in buffered peptone water and then selectively enriched in tetrathionate and Rappaport-Vassiliadis broths. Aliquots of the selective enrichment broths were combined for DNA extraction and analysis by the real-time PCR assay. When compared with the culture method, the diagnostic sensitivity of the PCR assay for the various matrices ranged from 97.1 to 100.0%, and the diagnostic specificity ranged from 91.3 to 100.0%. Kappa values ranged from 0.87 to 1.00, indicating excellent agreement of the real-time PCR assay to the culture method. The reduction in time and labor makes this highly sensitive and specific real-time PCR assay an excellent alternative to conventional culture methods for surveillance and research studies to improve food safety.

scholarly journals Probing the dissonance among the diagnostic outputs of multiple approaches used for detection of Methicillin-resistant Staphylococcus aureus (MRSA)

2020 ◽  
Author(s):  
Ujjwal Ranjan Dahiya ◽  
Arnab Sikidar ◽  
Priyanka Sharma ◽  
Chitra Rawat ◽  
Benu Dhawan ◽  
...  
Keyword(s):  
Staphylococcus Aureus ◽  
Methicillin Resistant Staphylococcus Aureus ◽  
Culture Method ◽  
Nucleic Acid Amplification ◽  
Detection Methods ◽  
Diagnostic Efficiency ◽  
Coagulase Negative Staphylococci ◽  
Culture Methods ◽  
Methicillin Resistant ◽  
Conventional Culture

Methicillin-resistant staphylococcus aureus (MRSA) is an extremely infectious hospital acquired bacterial pathogen often found in post-surgical patients globally. Early detection of such pathogens is a critical requirement to eliminate or reduce the incidence of antimicrobial resistance as well as for effective management of the disease. Despite the development of multiple biochemical, microbiological and nucleic acid amplification techniques (NAATs), conventional culture methods are widely used clinically owing to high variability between the methods, technical skills, and infrastructural needs. Further, multiple reports suggest a significant variation among diagnostic output for MRSA detection. This work attempts to probe the discordance among the diagnostic output of three commonly used methods while trying to understand the underlying cause of variability. MRSA detection on 217 clinical pus isolates was carried out using three different methods namely, conventional culture method, qPCR-based amplification, and a modern LAMP-based detection approach. Also, to confirm the presence of MRSA and distinguish from coagulase-negative staphylococci (CoNS), as well as to investigate the observed differences between qPCR and LAMP outputs, melt curve analysis was performed on discordant samples. LAMP-based MRSA detection was found to be the optimum method. In summary, this study evaluates the diagnostic efficiency of the different detection methods, while probing for possible explanations for the observed differences.

Evaluation of Detection Methods for Screening Meat and Poultry Products for the Presence of Foodborne Pathogens

2005 ◽  
Vol 68 (12) ◽  
pp. 2637-2647 ◽  
Author(s):  
VALERIE M. BOHAYCHUK ◽  
GARY E. GENSLER ◽  
ROBIN K. KING ◽  
JOHN T. WU ◽  
LYNN M. McMULLEN
Keyword(s):  
Foodborne Pathogens ◽  
Culture Method ◽  
Lateral Flow ◽  
Detection Methods ◽  
Enzyme Linked Immunosorbent Assay ◽  
Processed Meat ◽  
Culture Methods ◽  
E Coli ◽  
Poultry Products ◽  
Conventional Culture

Rapid and molecular technologies such as enzyme-linked immunosorbent assay (ELISA), PCR, and lateral flow immunoprecipitation can reduce the time and labor involved in screening food products for the presence of pathogens. These technologies were compared with conventional culture methodology for the detection of Salmonella, Campylobacter, Listeria, and Escherichia coli O157:H7 inoculated in raw and processed meat and poultry products. Recommended protocols were modified so that the same enrichment broths used in the culture methods were also used in the ELISA, PCR, and lateral flow immunoprecipitation assays. The percent agreement between the rapid technologies and culture methods ranged from 80 to 100% depending on the pathogen detected and the method used. ELISA, PCR, and lateral flow immunoprecipitation all performed well, with no statistical difference, compared with the culture method for the detection of E. coli O157:H7. ELISA performed better for the detection of Salmonella, with sensitivity and specificity rates of 100%. PCR performed better for the detection of Campylobacter jejuni, with 100% agreement to the culture method. PCR was highly sensitive for the detection of all the foodborne pathogens tested except Listeria monocytogenes. Although the lateral flow immunoprecipitation tests were statistically different from the culture methods for Salmonella and Listeria because of false-positive results, the tests did not produce any false negatives, indicating that this method would be suitable for screening meat and poultry products for these pathogens.

scholarly journals Rapid Detection and Identification Systems for the Microbiological Assessment of Processed Soy Foods: A Review

2020 ◽  
Vol 9 (5) ◽  
pp. 22
Author(s):  
Mitsuru Katase ◽  
Kazunobu Tsumura
Keyword(s):  
High Throughput ◽  
Culture Method ◽  
Detection Methods ◽  
Most Probable Number ◽  
Filter Method ◽  
Probable Number ◽  
Soy Foods ◽  
Flight Mass Spectrometry ◽  
Conventional Culture ◽  
Detection And Identification

Plant-based diets are gaining interest in promoting physical and environmental health worldwide. The widely growing consumption of processed soy foods results in an increased demand for safe and high quality soy foods. Many of the rapid bacterial detection methods currently available are inhibited by components in the food matrixes. In recent years, high-throughput devices have been developed, which aid in the enumeration and evaluation of microorganisms in processed soy foods (automated fluorescent filter method, high-throughput identification using matrix-assisted laser desorption ionization time-of-flight mass spectrometry, and automated most probable number method). These methods are more rapid and convenient compared to the conventional culture method. This review discusses alternate reliable methods for the microbiological assessment of processed soy foods, which guarantees the safety of the food delivered for consumption.

A randomized cross-over trial in patients suspected of PAD on diagnostic accuracy of ankle-brachial index by Doppler-based versus four-point oscillometry based measurements

VASA ◽  
2019 ◽  
Vol 48 (6) ◽  
pp. 516-522 ◽  
Author(s):  
Verena Mayr ◽  
Mirko Hirschl ◽  
Peter Klein-Weigel ◽  
Luka Girardi ◽  
Michael Kundi
Keyword(s):  
Diagnostic Accuracy ◽  
Ankle Brachial Index ◽  
Area Under The Curve ◽  
High Volume ◽  
Epidemiologic Studies ◽  
Lower Sensitivity ◽  
Oscillometric Device ◽  
Specificity And Sensitivity ◽  
Time Requirements ◽  
Peripheral Arterial

Summary. Background: For diagnosis of peripheral arterial occlusive disease (PAD), a Doppler-based ankle-brachial-index (dABI) is recommended as the first non-invasive measurement. Due to limitations of dABI, oscillometry might be used as an alternative. The aim of our study was to investigate whether a semi-automatic, four-point oscillometric device provides comparable diagnostic accuracy. Furthermore, time requirements and patient preferences were evaluated. Patients and methods: 286 patients were recruited for the study; 140 without and 146 with PAD. The Doppler-based (dABI) and oscillometric (oABI and pulse wave index – PWI) measurements were performed on the same day in a randomized cross-over design. Specificity and sensitivity against verified PAD diagnosis were computed and compared by McNemar tests. ROC analyses were performed and areas under the curve were compared by non-parametric methods. Results: oABI had significantly lower sensitivity (65.8%, 95% CI: 59.2%–71.9%) compared to dABI (87.3%, CI: 81.9–91.3%) but significantly higher specificity (79.7%, 74.7–83.9% vs. 67.0%, 61.3–72.2%). PWI had a comparable sensitivity to dABI. The combination of oABI and PWI had the highest sensitivity (88.8%, 85.7–91.4%). ROC analysis revealed that PWI had the largest area under the curve, but no significant differences between oABI and dABI were observed. Time requirement for oABI was significantly shorter by about 5 min and significantly more patients would prefer oABI for future testing. Conclusions: Semi-automatic oABI measurements using the AngER-device provide comparable diagnostic results to the conventional Doppler method while PWI performed best. The time saved by oscillometry could be important, especially in high volume centers and epidemiologic studies.

scholarly journals Evidence for the Use of Mucus Swabs to Detect Renibacterium salmoninarum in Brook Trout

Pathogens ◽  
2021 ◽  
Vol 10 (4) ◽  
pp. 460
Author(s):  
Tawni B. Riepe ◽  
Victoria Vincent ◽  
Vicki Milano ◽  
Eric R. Fetherman ◽  
Dana L. Winkelman
Keyword(s):  
Brook Trout ◽  
Sampling Methods ◽  
Kidney Tissue ◽  
Detection Methods ◽  
Fish Health ◽  
Wild Populations ◽  
Renibacterium Salmoninarum ◽  
Tissue Samples ◽  
Future Studies ◽  
Aquaculture Facilities

Efforts to advance fish health diagnostics have been highlighted in many studies to improve the detection of pathogens in aquaculture facilities and wild fish populations. Typically, the detection of a pathogen has required sacrificing fish; however, many hatcheries have valuable and sometimes irreplaceable broodstocks, and lethal sampling is undesirable. Therefore, the development of non-lethal detection methods is a high priority. The goal of our study was to compare non-lethal sampling methods with standardized lethal kidney tissue sampling that is used to detect Renibacterium salmoninarum infections in salmonids. We collected anal, buccal, and mucus swabs (non-lethal qPCR) and kidney tissue samples (lethal DFAT) from 72 adult brook trout (Salvelinus fontinalis) reared at the Colorado Parks and Wildlife Pitkin Brood Unit and tested each sample to assess R. salmoninarum infections. Standard kidney tissue detected R. salmoninarum 1.59 times more often than mucus swabs, compared to 10.43 and 13.16 times more often than buccal or anal swabs, respectively, indicating mucus swabs were the most effective and may be a useful non-lethal method. Our study highlights the potential of non-lethal mucus swabs to sample for R. salmoninarum and suggests future studies are needed to refine this technique for use in aquaculture facilities and wild populations of inland salmonids.

Detection by real-time quaking-induced conversion (RT-QuIC), ELISA, and IHC of chronic wasting disease prion in lymph nodes from Pennsylvania white-tailed deer with specific PRNP genotypes

2021 ◽  
pp. 104063872110214
Author(s):  
Deepanker Tewari ◽  
David Steward ◽  
Melinda Fasnacht ◽  
Julia Livengood
Keyword(s):  
Real Time ◽  
Disease Susceptibility ◽  
Chronic Wasting Disease ◽  
Low Frequency ◽  
The United States ◽  
Detection Methods ◽  
Spongiform Encephalopathy ◽  
Wasting Disease ◽  
Diagnostic Laboratory ◽  
Highly Sensitive

Chronic wasting disease (CWD) is a prion-mediated, transmissible disease of cervids, including deer ( Odocoileus spp.), which is characterized by spongiform encephalopathy and death of the prion-infected animals. Official surveillance in the United States using immunohistochemistry (IHC) and ELISA entails the laborious collection of lymphoid and/or brainstem tissue after death. New, highly sensitive prion detection methods, such as real-time quaking-induced conversion (RT-QuIC), have shown promise in detecting abnormal prions from both antemortem and postmortem specimens. We compared RT-QuIC with ELISA and IHC for CWD detection utilizing deer retropharyngeal lymph node (RLN) tissues in a diagnostic laboratory setting. The RLNs were collected postmortem from hunter-harvested animals. RT-QuIC showed 100% sensitivity and specificity for 50 deer RLN (35 positive by both IHC and ELISA, 15 negative) included in our study. All deer were also genotyped for PRNP polymorphism. Most deer were homozygous at codons 95, 96, 116, and 226 (QQ/GG/AA/QQ genotype, with frequency 0.86), which are the codons implicated in disease susceptibility. Heterozygosity was noticed in Pennsylvania deer, albeit at a very low frequency, for codons 95GS (0.06) and 96QH (0.08), but deer with these genotypes were still found to be CWD prion-infected.

scholarly journals Highly Sensitive Fluorescent Detection of Acetylcholine Based on the Enhanced Peroxidase-Like Activity of Histidine Coated Magnetic Nanoparticles

Nanomaterials ◽  
2021 ◽  
Vol 11 (5) ◽  
pp. 1207
Author(s):  
Hong Jae Cheon ◽  
Quynh Huong Nguyen ◽  
Moon Il Kim
Keyword(s):  
Magnetic Nanoparticles ◽  
High Sensitivity ◽  
High Specificity ◽  
Choline Oxidase ◽  
Fluorescent Detection ◽  
One Pot ◽  
Site Structure ◽  
Specificity And Sensitivity ◽  
Highly Sensitive ◽  
Peroxidase Mimics

Inspired by the active site structure of natural horseradish peroxidase having iron as a pivotal element with coordinated histidine residues, we have developed histidine coated magnetic nanoparticles (His@MNPs) with relatively uniform and small sizes (less than 10 nm) through one-pot heat treatment. In comparison to pristine MNPs and other amino acid coated MNPs, His@MNPs exhibited a considerably enhanced peroxidase-imitating activity, approaching 10-fold higher in catalytic reactions. With the high activity, His@MNPs then were exploited to detect the important neurotransmitter acetylcholine. By coupling choline oxidase and acetylcholine esterase with His@MNPs as peroxidase mimics, target choline and acetylcholine were successfully detected via fluorescent mode with high specificity and sensitivity with the limits of detection down to 200 and 100 nM, respectively. The diagnostic capability of the method is demonstrated by analyzing acetylcholine in human blood serum. This study thus demonstrates the potential of utilizing His@MNPs as peroxidase-mimicking nanozymes for detecting important biological and clinical targets with high sensitivity and reliability.

Comparison of a Real-Time PCR Method with a Culture Method for the Detection of Salmonella enterica Serotype Enteritidis in Naturally Contaminated Environmental Samples from Integrated Poultry Houses

2012 ◽  
Vol 75 (4) ◽  
pp. 743-747 ◽  
Author(s):  
BWALYA LUNGU ◽  
W. DOUGLAS WALTMAN ◽  
ROY D. BERGHAUS ◽  
CHARLES L. HOFACRE
Keyword(s):  
Real Time ◽  
Real Time Pcr ◽  
Salmonella Enteritidis ◽  
Culture Method ◽  
Pcr Assay ◽  
Culture Methods ◽  
The Real ◽  
Selective Enrichment ◽  
Conventional Culture ◽  
Field Samples

Conventional culture methods have traditionally been considered the “gold standard” for the isolation and identification of foodborne bacterial pathogens. However, culture methods are labor-intensive and time-consuming. A Salmonella enterica serotype Enteritidis–specific real-time PCR assay that recently received interim approval by the National Poultry Improvement Plan for the detection of Salmonella Enteritidis was evaluated against a culture method that had also received interim National Poultry Improvement Plan approval for the analysis of environmental samples from integrated poultry houses. The method was validated with 422 field samples collected by either the boot sock or drag swab method. The samples were cultured by selective enrichment in tetrathionate broth followed by transfer onto a modified semisolid Rappaport-Vassiliadis medium and then plating onto brilliant green with novobiocin and xylose lysine brilliant Tergitol 4 plates. One-milliliter aliquots of the selective enrichment broths from each sample were collected for DNA extraction by the commercial PrepSEQ nucleic acid extraction assay and analysis by the Salmonella Enteritidis–specific real-time PCR assay. The real-time PCR assay detected no significant differences between the boot sock and drag swab samples. In contrast, the culture method detected a significantly higher number of positive samples from boot socks. The diagnostic sensitivity of the real-time PCR assay for the field samples was significantly higher than that of the culture method. The kappa value obtained was 0.46, indicating moderate agreement between the real-time PCR assay and the culture method. In addition, the real-time PCR method had a turnaround time of 2 days compared with 4 to 8 days for the culture method. The higher sensitivity as well as the reduction in time and labor makes this real-time PCR assay an excellent alternative to conventional culture methods for diagnostic purposes, surveillance, and research studies to improve food safety.

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