scholarly journals Validation of a Commercial Loop-Mediated Isothermal Amplification (LAMP)-Based Kit for the Detection of Salmonella spp. According to ISO 16140:2016

2021 ◽  
Vol 11 (15) ◽  
pp. 6669
Author(s):  
Calogero Di Bella ◽  
Antonella Costa ◽  
Sonia Sciortino ◽  
Giuseppa Oliveri ◽  
Gaetano Cammilleri ◽  
...  
Keyword(s):  
Real Time ◽  
Real Time Pcr ◽  
Reference Method ◽  
Poultry Meat ◽  
Lamp Method ◽  
Salmonella Spp ◽  
Cultural Method ◽  
Detection And Identification ◽  
Biochemical Identification ◽  
Raw Poultry

The traditional cultural method (PCR and Real-Time PCR) for Salmonella spp. detection and identification is laborious and time-consuming. A qualitative LAMP method detecting Salmonella spp. was validated in compliance with ISO 16140:2016. The results show a relative accuracy, sensitivity, and specificity of 100% in comparison with the reference method ISO 6579-1:2017; the LOD50 was set as 0.4 CFU/g. Additionally, a field study was carried out comparing the LAMP kit, a commercially available Real-Time PCR kit (FoodProof Salmonella, Biotecon Diagnostics), and the reference cultural method. The Salmonella spp. LAMP kit was suitable for reliable detection of Salmonella spp., simplifying and reducing the extent and the steps of the analytical process. A total of 105 samples of raw poultry meat were screened for the presence of Salmonella spp. according to three methods: the LAMP kit Salmonella spp. (Enbiotech), the Real-Time PCR kit FoodProof Salmonella (Biotecon), and the reference cultural method. Using these three methods, only one sample out of the 105 (0.95%) tested was positive for Salmonella spp. This sample was further investigated using the reference method described in ISO 6579-3:2014, in order to characterise the Salmonella strain. Following this further biochemical identification and serological typing, the isolate was characterised as Salmonella Infantis.

scholarly journals Evaluation of Applied Biosystems MicroSEQ® Real-Time PCR System for Detection of Salmonella spp. in Food

2011 ◽  
Vol 94 (4) ◽  
pp. 1106-1116 ◽  
Author(s):  
Priya Balachandran ◽  
Yanxiang Cao ◽  
Lily Wong ◽  
Manohar R Furtado ◽  
Olga V Petrauskene ◽  
...  
Keyword(s):  
Real Time ◽  
Study Design ◽  
Real Time Pcr ◽  
Detection System ◽  
Peanut Butter ◽  
Reference Method ◽  
Black Pepper ◽  
Salmonella Spp ◽  
Culture Methods ◽  
Significant Difference

Abstract Real-time PCR methods for detecting foodborne pathogens offer the advantages of simplicity and quick time-to-results compared to traditional culture methods. In this study, the MicroSEQ® real-time PCR system was evaluated for detection of Salmonella spp. in 10 different food matrixes following the AOAC Research Institute's Performance Tested MethodSM validation program. In addition, the performance of the MicroSEQ system was evaluated for the detection of Salmonella in peanut butter as a part of the Emergency Response Validation Program sponsored by the AOAC Research Institute. The system was compared to the ISO 6579 reference method using a paired-study design for detecting Salmonella spp. in raw ground beef, raw chicken, raw shrimp, Brie cheese, shell eggs, cantaloupe, chocolate, black pepper, dry infant formula, and dry pet food. For the peanut butter study, the system was compared to the U.S. Food and Drug Administration's Bacteriological Analytical Manual procedures using an unpaired-study design. No significant difference in performance was observed between the MicroSEQ Salmonella spp. detection system and the corresponding reference methods for all 11 food matrixes. The MicroSEQ system detected all Salmonella strains tested, while showing good discrimination against detection of an exclusivity panel of 30 strains, with high accuracy.

MicroSEQ® Salmonella spp. Detection Kit Using the Pathatrix® 10-Pooling Salmonella spp. Kit Linked Protocol Method Modification

2014 ◽  
Vol 97 (2) ◽  
pp. 484-491 ◽  
Author(s):  
Jason Wall ◽  
Rick Conrad ◽  
Kathy Latham ◽  
Eric Liu
Keyword(s):  
Real Time ◽  
Study Design ◽  
Foodborne Pathogens ◽  
Real Time Pcr ◽  
Reference Method ◽  
Immunomagnetic Separation ◽  
Pcr Analysis ◽  
Salmonella Spp ◽  
Culture Methods ◽  
Design For All

Abstract Real-time PCR methods for detecting foodborne pathogens offer the advantages of simplicity and quick time to results compared to traditional culture methods. The addition of a recirculating pooled immunomagnetic separation method prior to real-time PCR analysis increases processing output while reducing bothcost and labor. This AOAC Research Institute method modification study validates the MicroSEQ®Salmonella spp. Detection Kit [AOAC Performance Tested Method(PTM) 031001] linked with the Pathatrix® 10-Pooling Salmonella spp. Kit (AOAC PTM 090203C) in diced tomatoes, chocolate, and deli ham. The Pathatrix 10-Pooling protocol represents a method modification of the enrichment portion of the MicroSEQ Salmonella spp. protocol. The results of the method modification were compared to standard cultural reference methods for diced tomatoes, chocolate, and deli ham. All threematrixes were analyzed in a paired study design. An additional set of chocolate test portions was analyzed using an alternative enrichment medium in an unpaired study design. For all matrixes tested, there were no statistically significant differences in the number of positive test portions detected by the modified candidate method compared to the appropriate reference method. The MicroSEQ Salmonella spp. protocol linked with the Pathatrix individual or 10-Pooling procedure demonstrated reliabilityas a rapid, simplified, method for the preparation of samples and subsequent detection of Salmonella in diced tomatoes, chocolate, and deliham.

scholarly journals Application of a Real-Time PCR Method for Salmonella spp., Escherichia coli, Staphylococcus aureus and Clostridium perfringens Detection in Water Samples

2013 ◽  
Vol 62 (4) ◽  
pp. 439-443
Author(s):  
BEATA ROZWADOWSKA ◽  
MARTA ALBERTYŃSKA ◽  
GRZEGORZ HUDZIK ◽  
HUBERT OKŁA ◽  
KRZYSZTOF P. JASIK ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Staphylococcus Aureus ◽  
Real Time ◽  
Clostridium Perfringens ◽  
Water Samples ◽  
Real Time Pcr ◽  
Culture Method ◽  
Salmonella Spp ◽  
Detection And Identification ◽  
Pcr Method

The diagnostic assessment of water sanitary state is based mainly on the cultivation of bacteria retained on membrane filters. However classical microbiology methods have a lot of disadvantages. More and more frequently, rapid detection and identification of pathogens present in water is based on molecular biology techniques. The aim of this study was to determine the effectiveness and usefulness of a real-time PCR method, when compared to the recommended bacteria culture method, in diagnostics of pathogens in water samples. The research concerned the detection and identification of main sanitary indicators of water such as: Salmonella spp., Escherichia coli, Staphylococcus aureus and Clostridium perfringens. The analyses were conducted in water samples contaminated with the reference material (the aforementioned bacteria) and real environmental samples, which were examined for the presence of nucleic acid of: Salmonella spp., E. coli, S. aureus and C. perfringens using a real-time PCR method.

scholarly journals A single-run, real-time PCR for detection and identification ofBorrelia burgdorferi sensu latospecies, based on thehbbgene sequence

2006 ◽  
Vol 259 (1) ◽  
pp. 35-40 ◽  
Author(s):  
Denis Portnoï ◽  
Natacha Sertour ◽  
Elisabeth Ferquel ◽  
Martine Garnier ◽  
Guy Baranton ◽  
...  
Keyword(s):  
Real Time ◽  
Real Time Pcr ◽  
Detection And Identification

Development of a single-tube duplex EvaGreen real-time PCR for the detection and identification of EHV-1 and EHV-4

2014 ◽  
Vol 98 (9) ◽  
pp. 4179-4186 ◽  
Author(s):  
Zhe Hu ◽  
Chao Zhu ◽  
Hao Chang ◽  
Wei Guo ◽  
Diqiu Liu ◽  
...  
Keyword(s):  
Real Time ◽  
Real Time Pcr ◽  
Single Tube ◽  
Detection And Identification

scholarly journals Primers designed for the detection of grapevine pathogens spreading with propagating material by quantitative real-time PCR

2015 ◽  
Vol 21 (1-2) ◽  
Author(s):  
N. Czotter ◽  
E. Manduláné Farkas ◽  
R. Lózsa ◽  
I. Ember ◽  
G. Szûcsné Varga ◽  
...  
Keyword(s):  
Real Time ◽  
Real Time Pcr ◽  
Dna Amplification ◽  
Molecular Techniques ◽  
Latent Infections ◽  
Quantitative Real Time Pcr ◽  
Detection And Identification

Several grapevine pathogens are disseminated by propagating material as systemic, but latent infections. Their detection and identification have a basic importance in the production and handling of propagating stocks. Thus several sensitive and reliable diagnostic protocols mostly based on molecular techniques have been developed. Of these methods quantitative real-time PCR (q-PCR) has recently got an emerging importance. Here we collected primer data for the detection and identification of grapevine pathogens which are important in the production of propagating stocks by q-PCR. Additional novel techniques that use DNA amplification, hybridization and  sequencing are also briefly reviewed.

scholarly journals Rapid Detection and Identification of Uveitis Pathogens by Qualitative Multiplex Real-Time PCR

2018 ◽  
Vol 59 (1) ◽  
pp. 582 ◽  
Author(s):  
Paulo J. M. Bispo ◽  
Samaneh Davoudi ◽  
Matthew L. Sahm ◽  
Ai Ren ◽  
John Miller ◽  
...  
Keyword(s):  
Real Time ◽  
Real Time Pcr ◽  
Rapid Detection ◽  
Detection And Identification

scholarly journals Rapid identification of transgenic cotton (Gossypium hirsutum L.) plants by loop-mediated isothermal amplification

2011 ◽  
Vol 47 (No. 4) ◽  
pp. 140-148 ◽  
Author(s):  
N. Rostamkhani ◽  
A. Haghnazari ◽  
M. Tohidfar ◽  
A. Moradi
Keyword(s):  
Real Time ◽  
Real Time Pcr ◽  
Lamp Assay ◽  
Transgenic Cotton ◽  
Rapid Identification ◽  
Isothermal Amplification ◽  
Detection Sensitivity ◽  
Lamp Method ◽  
Loop Mediated Isothermal Amplification ◽  
Leaf Tissues

In an attempt to speed up the process of screening of transgenic cotton (G. hirsutum L.) plants, a visual and rapid loop-mediated isothermal amplification (LAMP) assay was adopted. Genomic DNA was extracted from fresh leaf tissues of T<sub>2</sub> transgenic cotton containing chitinase (chi) and cry1A(b) genes. Detection of genes of interest was performed by polymerase chain reaction (PCR), LAMP and real-time PCR methods. In LAMP assay the amplification was performed after 30 min at 65&deg;C when loop primers were involved in the reaction. The involvement of loop primers decreased the time needed for amplification. By testing serial tenfold dilutions (10<sup>&ndash;1</sup> to 10<sup>&ndash;8</sup>) of the genes of interest, the detection sensitivity of LAMP was found to be 100-fold higher than that of PCR. The rapid DNA extraction method and LAMP assay can be performed within 30 min and the derived LAMP products can be directly observed as visually detectable based on turbidity in the reaction tube. The accuracy of LAMP method in the screening of transgenes was confirmed by PCR and real-time PCR. The developed method was efficient, rapid and sensitive in the screening of cotton transgenic plants. This method can be applied to any other crops.

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