scholarly journals A new method for rapid detection of Listeria monocytogenes in food

2000 ◽  
Vol 18 (No. 3) ◽  
pp. 103-109 ◽  
Author(s):  
K. Zdeňková ◽  
K. Demnerová ◽  
G. Jeníková ◽  
J. Pazlarová
Keyword(s):  
Listeria Monocytogenes ◽  
Pcr Primers ◽  
Immunomagnetic Separation ◽  
Food Samples ◽  
Special Importance ◽  
Ground Meat ◽  
Specific Detection ◽  
Gene Encoding ◽  
Pcr Method

Listeria monocytogenes represents serious danger for human health. Thus detection of this pathogen in food, which represents its main means of entry into the organism, is a topic of special importance. The original classic methods for the determination of Listeria monocytogenes are in general laborious and time-consuming procedures. In order to address this issue we developed a new rapid method for specific detection of Listeria monocytogenes in food samples. The method consists of three steps: i) enrichment of food microflora (16 h), ii) selective isolation of Listeria sp. exploiting immunomagnetic separation (2–3 h) followed by iii) precise identification of Listeria sp. and Listeria monocytogenes using duplex PCR. PCR primers specific to part of 16S rRNA were used in order to identify the members of Listeria genus. The specific identification of Listeria monocytogenes was accomplished exploiting a pair of primers specific to gene encoding invasion-associated protein – iap (4–5 h). Amplification products, 1003 bp and 593 bp respectively, were separated by electrophoresis and visualized by UV detection. The optimized IMS-PCR method was used to test the presence of Listeria sp. and Listeria monocytogenes in food samples (ground meat, low-fat milk and cheese [olomoucké tvarůžky]). A comparison of the efficiency of the bacteria enrichment step by IMS and centrifugation was also performed. The analysis time including enrichment is less than 24 h. The detection limit for Listeria monocytogenes was found between 101–102 cfu per 25 g of food sample.

Combination of Immunomagnetic Separation and Polymerase Chain Reaction for the Simultaneous Detection of Listeria monocytogenes and Salmonella spp. in Food Samples

2001 ◽  
Vol 64 (11) ◽  
pp. 1744-1750 ◽  
Author(s):  
HSIEN-YEE HSIH ◽  
HAU-YANG TSEN
Keyword(s):  
Polymerase Chain Reaction ◽  
Listeria Monocytogenes ◽  
Simultaneous Detection ◽  
Meat Products ◽  
Immunomagnetic Separation ◽  
Food Samples ◽  
Chain Reaction ◽  
Salmonella Spp ◽  
Pcr Method ◽  
Polymerase Chain

A method that combined the immunomagnetic separation (IMS) technique and the multiplex polymerase chain reaction (PCR) method (i.e., the IMS-mPCR method) was developed for simultaneous detection of Listeria monocytogenes and Salmonella spp. in food samples. When only the multiplex PCR method was used, it was found that if cell numbers of each of the two target organisms (L. monocytogenes and Salmonella spp.) were above the detection limit, but differed by more than 2 logs—e.g., n × 107 to n × 104 or n × 106 to n × 103—the organism presenting the lower numbers might go undetected. Following the enrichment step with universal preenrichment (UP) broth, if an IMS method using equal quantities of anti-Listeria and anti-Salmonella immunomagnetic beads was performed prior to PCR, both pathogens could be detected unambiguously. Such results could be obtained for target organisms in food samples, such as milk, dairy, and meat products, if similar enrichment and IMS steps were performed prior to PCR.

scholarly journals Optimization of a Viability PCR Method for the Detection of Listeria monocytogenes in Food Samples

2018 ◽  
Vol 75 (6) ◽  
pp. 779-785 ◽  
Author(s):  
Gemma Agustí ◽  
Mariana Fittipaldi ◽  
Francesc Codony
Keyword(s):  
Listeria Monocytogenes ◽  
Food Samples ◽  
Pcr Method

scholarly journals Gas Gangrene of Different Origin Associated with Clostridium perfringens Type A in Three Patients Simultaneously Hospitalized in a Single Department of Orthopedics and Traumatology in Poland

2016 ◽  
Vol 65 (4) ◽  
pp. 399-406
Author(s):  
Monika Brzychczy-Włoch ◽  
Dorota Ochońska ◽  
Anna Piotrowska ◽  
Małgorzata Bulanda
Keyword(s):  
Clostridium Perfringens ◽  
Gas Gangrene ◽  
Pulsed Field Gel Electrophoresis ◽  
Gene Encoding ◽  
The Third ◽  
Resistance Patterns ◽  
Biochemical Profiles ◽  
Pcr Method ◽  
Encoding Genes

The objective of the study was to perform a comparative analysis of phenotypic and genetic similarity, determination of resistance profiles, detection of toxin-encoding genes and molecular typing of Clostridium perfringens isolates originating from patients with gas gangrene. The study encompassed three patients with a clinical and microbiological diagnosis of gas gangrene who were hospitalized in one of the hospitals of the Kujawsko-Pomorskie province in the same period of time between 8th April 2015 and 20th April 2015. The three C. perfringens isolates studied had identical biochemical profiles. Two isolates had identical resistance patterns, while the third presented a different profile. Using the multiplex PCR method, all isolates showed the presence of cpa gene encoding α-toxin; furthermore, the presence of the cpb2 gene encoding β2-toxin was confirmed in two isolates. Genotyping with the use of pulsed field gel electrophoresis (PFGE) indicated that the isolates originating from the three studied patients represent three genetically different restrictive patterns which corresponded to three different clones – clone A, clone B and clone C. As a result of the study, it is possible to conclude that the studied patients simultaneously hospitalized in a single Department of Orthopedics and Traumatology developed three different endogenous infections.

scholarly journals Rapid Determination of Listeria monocytogenes by Automated Enzyme-Linked Immunoassay and Nonradioactive DNA Probe

2000 ◽  
Vol 83 (1) ◽  
pp. 86-88 ◽  
Author(s):  
Khalil F Kerdahi ◽  
Philip F Istafanos
Keyword(s):  
Listeria Monocytogenes ◽  
Rapid Determination ◽  
Food Samples ◽  
Dna Probe ◽  
Negative Results ◽  
Smoked Salmon ◽  
Colony Forming Units ◽  
Low Levels ◽  
Analytical Time

Abstract A rapid and reliable analytical method was developed to detect and confirm the presence of Listeria monocytogenes in raw and partially processed foods. Forty-nine food samples (25 mixed cut vegetable salad, 12 smoked salmon, and 12 sterile smoked salmon) were individually inoculated with high levels [10–100 colony forming units (cfu)/25 g sample] and low levels (1–10 cfu/25 g sample) of L. monocytogenes, and were screened using the Vitek Immuno Diagnostic Assay (VIDAS) Listeria monocytogenes (VIDAS LMO)]. Positive test results were confirmed as L. monocytogenes by nonradioactive DNA probe. All samples inoculated with high levels of L. monocytogenes were detected by VIDAS and 96% were confirmed as L. monocytogenes by DNA probe. VIDAS LMO detected 89% of samples inoculated with low levels of L. monocytogenes, and 87% of these were confirmed as positive by DNA probe. In addition, 12 other samples (4 from each of mixed cut vegetable salad, smoked salmon, and sterile smoked salmon) were inoculated with high levels of L. ivanovii, L. seeligeri, L. welshimeri, L. innocua, L. grayi, and L. murrayi. Samples were assayed by the same protocol and all gave negative results. Compared with the cultural method, the VIDAS LMO nonradioactive DNA probe combination is highly specific, discriminates between L. monocytogenes and all other Listeria species, and reduces analytical time.

Development and Use of Polymerase Chain Reaction for the Specific Detection of Salmonella Typhimurium in Stool and Food Samples

1999 ◽  
Vol 62 (10) ◽  
pp. 1103-1110 ◽  
Author(s):  
JER-SHENG LIN ◽  
HAU-YANG TSEN
Keyword(s):  
Polymerase Chain Reaction ◽  
Salmonella Typhimurium ◽  
Pcr Primers ◽  
Food Samples ◽  
Clinical Samples ◽  
Specific Detection ◽  
Chain Reaction ◽  
Pcr Product ◽  
Polymerase Chain ◽  
Salmonella Serovars

Salmonella Typhimurium is one of the most important Salmonella serovars that may cause foodborne disease and human salmonellosis infection. Detection of this organism in the clinical samples of persons with gastroenteritis and the food samples associated with such persons may allow us to trace the cause of disease. Because malic acid dehydrogenase, an enzyme of the citric acid cycle, is common to organisms, the gene (mdh) coding for this enzyme was selected for the design of Salmonella Typhimurium-specific polymerase chain reaction (PCR) primers. By comparison of the mdh gene sequences of Salmonella Typhimurium and other Salmonella serotypes and of some isolates of other genera, two oligonucleotides were designed and used as PCR primers for the specific detection of Salmonella Typhimurium. The molecular weight of the PCR product was 261 bp as expected. Salmonella serovars other than Salmonella Typhimurium and isolates of other genera in the Enterobacteriaceae that is closely related to Salmonella did not generate any false-positive results. When this primer pair was used for the detection of Salmonella Typhimurium cells artificially inoculated into human stool specimens and food samples, such as milk and raw chicken meat, levels as low as 100 CFU per 0.1 g of stool specimen or per ml of milk or food homogenate could be detected if an 8- to 12-h preculture step using combined lactose-tetrathionate broth was performed prior to the PCR.

scholarly journals Determination of Listeria monocytogenes numbers at less than 10 cfu/g

2017 ◽  
Vol 56 (1) ◽  
pp. 25-30 ◽  
Author(s):  
K. Hunt ◽  
M. Vacelet ◽  
K. Jordan
Keyword(s):  
Listeria Monocytogenes ◽  
Detection Limit ◽  
Agar Plate ◽  
Foodborne Pathogen ◽  
Petri Dish ◽  
Food Samples ◽  
Plate Method ◽  
Pure Cultures ◽  
Response Current

AbstractListeria monocytogenes is a foodborne pathogen that causes a relatively rare foodborne disease called listeriosis, with a high mortality rate of 20%-30% and an undefined dose response. Current European Union regulations permit up to 100 colony-forming units (cfu)/g in food at the end of its shelf life, where the food has been shown not to support the growth of this pathogenic bacterium. Therefore, enumeration of L. monocytogenes at low numbers in food is important. The objective of this study was to reduce the detection limit of L. monocytogenes in food by a factor of 10. The International Organisation for Standardisation (ISO) 11290-2 method for enumeration of L. monocytogenes in food recommends spreading 0.1 mL of a 1:10 dilution of the food on the surface of an agar plate (detection limit 100 cfu/g), or 1.0 mL spread in equal parts on the surface of three agar plates (detection limit: 10 cfu/g). The pour-plate method (using 1 or 10 mL of an appropriate dilution) was compared to the spread-plate method using the ISO-approved chromogenic medium Agar Listeria according to Ottaviani and Agosti (ALOA). Using the pour-plate method, the colony morphology and halo formation were similar to the spread-plate method from pure cultures and inoculated foods. Using the pour-plate method in a 140 mm Petri dish, 10 mL of a 1:10 dilution of food allowed determination of numbers as low as 1 cfu/g. Applying this method, L. monocytogenes in naturally contaminated food samples were enumerated at numbers as low as 1-9 cfu/g.

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