Direct Detection and Typing of Variety Pathogens Important for Human and Animal Health Using xMAP Technology

Speaker
  • Head of Department of Food and Feed Safety, Veterinary Research Institute

Abstract

Background and aim:
Rapid and complex diagnosis of viral, bacterial and parasitic infections is highly desirable not only in clinical practice, but also in food safety or early warning systems. Multiple Oligonucleotide Ligation followed by
PCR (MOL-PCR) is easy to set up and allows to simply add detection targets to the existing panels. The MOL-PCR technique was used in the development of the panels for the detection and identification of viruses and bacteria originating from food and feed, viral pathogens of swine, bacterial pathogens in small ruminants and biothreat agents.

Methods:
MOL-PCR is based on the ligation of specific oligonucleotides in the presence of a specific target DNA. In the case of RNA viruses, reverse transcription is required. The ligation product serves as a template for the amplification with universal primers, one of which is labelled, in subsequent PCR. Visualization of PCR products is mediated by binding to magnetic fluorescent beads in a suspension array and is performed on a
MAGPIX® (Luminex) instrument. Each panel contained its own internal amplification control, which served to the differentiation of truly negative and false negative samples and control of presence of inhibitors. All the panels underwent the validation procedure adopted from the guidelines for the validation of the qPCR assays developed by FDA.

Results:
The result is a total of 5 validated panels for rapid detection and identification of the desired agents. The first includes viruses that are associated with food and water contamination - noroviruses, hepatitis A and E virus, rotaviruses and adenoviruses. The second panel is able to detect food-derived bacteria - Salmonella, Yersinia, Cronobacter, or pathogenic E. coli. Next panel is designed to detect viral agents in pigs such as African swine fever or various types of influenza. Panel dedicated to the small ruminants covers M. a. subsp. paratuberculosis, C. pseudotuberculosis, M. bovis and M. tuberculosis complex members - agents responsible for the tuberculosis and pseudotuberculosis infections. The last panel is developed for the needs of security forces and detects and typifies biothreat agents -B. anthracis, Y. pestis, F. tularensis and Brucella spp.

Conclusion:
The developed panels have been validated for complex diagnostics of clinical material, food including water, animals and other relevant matrices. The process of optimization of MOL-PCR was established and critical points, which require a special attention in the optimization were identified. Development of all the panels was targeted with the intention to fulfill the demands placed on the diagnostic tools.

 


Show Resources
You May Also Like
SEP 17, 2021 12:00 PM CST
C.E. CREDITS
SEP 17, 2021 12:00 PM CST
Date: September 16, 2021 Time: 9:00pm (PDT), 12:00am (EDT) 3D cellular models like organoids and spheroids offer an opportunity to better understand complex biology in a physiologically rele...
APR 21, 2021 10:00 AM PDT
C.E. CREDITS
APR 21, 2021 10:00 AM PDT
Date: April 21, 2021 Time: 10:00am (PDT), 1:00pm (EDT) Serological testing for SARS-CoV-2 has been steadily adopted into clinical practice over the course of this pandemic. In this webinar,...
SEP 14, 2021 7:00 AM PDT
C.E. CREDITS
SEP 14, 2021 7:00 AM PDT
Date: September 14, 2021 Time: 7am PDT, 10am EDT, 4pm CEST A conventional thermal cycler has long been a commodity product in the lab and end-point PCR techniques can be completed almost wit...
MAY 18, 2021 8:00 AM PDT
C.E. CREDITS
MAY 18, 2021 8:00 AM PDT
Date: May 18, 2021 Time: 8:00am (PST) The global pandemic has caused an increased focus and scrutiny on molecular diagnostic assay development, resulting in a need for assays that provide qu...
APR 01, 2021 8:00 AM PDT
C.E. CREDITS
APR 01, 2021 8:00 AM PDT
Date: April 01, 2021 Time: 8:00am (PST), 11:00am (EST) Generating therapeutic antibodies is far more challenging than obtaining antibodies that merely recognize their targets. Engineering po...
JUN 03, 2021 12:00 PM CST
JUN 03, 2021 12:00 PM CST
DATE: June 3, 2021 TIME: 12:00pm SGT This webinar is a virtual event that focuses on utilizing the Gibco CTS Rotea System for Cell and Gene Therapy...

Direct Detection and Typing of Variety Pathogens Important for Human and Animal Health Using xMAP Technology


No demographic data is available yet for this event.


Show Resources
Loading Comments...
Show Resources