Development and Single Laboratory Evaluation of a Refined and specific Real-time PCR Detection Method, Using Mitochondrial Primers (Mit1C), for the Detection of Cyclospora cayetanensis in Produce.

Animals Cyclospora / genetics Real-Time Polymerase Chain Reaction / methods Oocysts Rubus Coriandrum Cyclosporiasis / diagnosis
Cyclospora cayetanensis Foodborne Leafy greens Mitochondria Parasite Real-time PCR

Journal

Journal of food protection
ISSN: 1944-9097
Titre abrégé: J Food Prot
Pays: United States
ID NLM: 7703944

Informations de publication

Date de publication:
02 2023
Historique:
received: 03 10 2022
revised: 20 12 2022
accepted: 29 12 2022
entrez: 14 3 2023
pubmed: 15 3 2023
medline: 16 3 2023
Statut: ppublish

Résumé

Regulatory methods for detection of the foodborne protozoan parasite Cyclospora cayetanensis must be specific and sensitive. To that end, we designed and evaluated (in a single laboratory validation) a novel and improved primer/probe combination (Mit1C) for real-time PCR detection of C. cayetanensis in produce. The newly developed primer/probe combination targets a conserved region of the mitochondrial genome of C. cayetanensis that varies in other closely related organisms. The primer/probe combination was evaluated both in silico and using several real-time PCR kits and polymerases against an inclusivity/exclusivity panel comprised of a variety of C. cayetanensis oocysts, as well as DNA from other related Cyclospora spp. and closely related parasites. The new primer/probe combination amplified only C. cayetanensis, thus demonstrating specificity. Sensitivity was evaluated by artificially contaminating cilantro, raspberries, and romaine lettuce with variable numbers (200 and 5) of C. cayetanensis oocysts. As few as 5 oocysts were detected in 75%, 67.7%, and 50% of the spiked produce samples (cilantro, raspberries, and romaine lettuce), respectively, all uninoculated samples and no-template real-time PCR controls were negative. The improved primer/probe combination should prove an effective analytical tool for the specific detection of C. cayetanensis in produce.

Identifiants

DOI: 10.1016/j.jfp.2022.100037 PMID: 36916572
pubmed: 36916572
pii: S0362-028X(22)11952-6
doi: 10.1016/j.jfp.2022.100037
pii:
doi:

Types de publication

Journal Article Research Support, U.S. Gov't, P.H.S.

Langues

eng

Sous-ensembles de citation

IM

Pagination

100037

Informations de copyright

Published by Elsevier Inc.

Déclaration de conflit d'intérêts

Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper.

Auteurs

Kannan V Balan (KV)

U.S. Food and Drug Administration, Center for Food Safety and Applied Nutrition, Office of Applied Research and Safety Assessment, Division of Virulence Assessment, Laurel, MD 20708, USA.

Mark Mammel (M)

U.S. Food and Drug Administration, Center for Food Safety and Applied Nutrition, Office of Applied Research and Safety Assessment, Division of Molecular Biology, Laurel, MD 20708, USA.

David Lipman (D)

U.S. Food and Drug Administration, Center for Food Safety and Applied Nutrition, Office of the Center Director, College Park, MD 20740, USA.

Uma Babu (U)

U.S. Food and Drug Administration, Center for Food Safety and Applied Nutrition, Office of Applied Research and Safety Assessment, Division of Virulence Assessment, Laurel, MD 20708, USA.

Lisa M Harrison (LM)

U.S. Food and Drug Administration, Center for Food Safety and Applied Nutrition, Office of Applied Research and Safety Assessment, Division of Virulence Assessment, Laurel, MD 20708, USA.

Sonia Almeria (S)

U.S. Food and Drug Administration, Center for Food Safety and Applied Nutrition, Office of Applied Research and Safety Assessment, Division of Virulence Assessment, Laurel, MD 20708, USA.

Mauricio Durigan (M)

U.S. Food and Drug Administration, Center for Food Safety and Applied Nutrition, Office of Applied Research and Safety Assessment, Division of Virulence Assessment, Laurel, MD 20708, USA.

Susan R Leonard (SR)

U.S. Food and Drug Administration, Center for Food Safety and Applied Nutrition, Office of Applied Research and Safety Assessment, Division of Molecular Biology, Laurel, MD 20708, USA.

Hyein Jang (H)

U.S. Food and Drug Administration, Center for Food Safety and Applied Nutrition, Office of Applied Research and Safety Assessment, Division of Virulence Assessment, Laurel, MD 20708, USA.

Solomon Gebru (S)

U.S. Food and Drug Administration, Center for Food Safety and Applied Nutrition, Office of Applied Research and Safety Assessment, Division of Virulence Assessment, Laurel, MD 20708, USA.

John Grocholl (J)

U.S. Food and Drug Administration, Center for Food Safety and Applied Nutrition, Office of Applied Research and Safety Assessment, Division of Virulence Assessment, Laurel, MD 20708, USA.

Socrates Trujillo (S)

U.S. Food and Drug Administration, Center for Food Safety and Applied Nutrition, Office of Applied Research and Safety Assessment, Division of Virulence Assessment, Laurel, MD 20708, USA.

Kelli L Hiett (KL)

U.S. Food and Drug Administration, Center for Food Safety and Applied Nutrition, Office of Applied Research and Safety Assessment, Office of the Director, Laurel, MD 20708, USA. Electronic address: Kelli.Hiett@FDA.HHS.GOV.

Steve Musser (S)

U.S. Food and Drug Administration, Center for Food Safety and Applied Nutrition, Office of the Center Director, College Park, MD 20740, USA.

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Classifications MeSH

questionsmedicales.fr › Eucaryotes › Animaux › Development and Single Laboratory Evaluation...
questionsmedicales.fr › Eucaryotes › Alveolata › Apicomplexa › Coccidia › Eimeriida › Eimeriidae › Cyclospora › Development and Single Laboratory Evaluation...
questionsmedicales.fr › Techniques d'investigation › Techniques génétiques › Techniques d'amplification d'acides nucléiques › Réaction de polymérisation en chaîne › Réaction de polymérisation en chaine en temps réel › Development and Single Laboratory Evaluation...
questionsmedicales.fr › Phénomènes physiologiques › Croissance et développement › Morphogenèse › Métamorphose biologique › Étapes du cycle de vie › Oocystes › Development and Single Laboratory Evaluation...
questionsmedicales.fr › Eucaryotes › Viridiplantae › Streptophyta › Embryophyta › Tracheobionta › Magnoliopsida › Rosanae › Rosales › Rosaceae › Rubus › Development and Single Laboratory Evaluation...
questionsmedicales.fr › Eucaryotes › Viridiplantae › Streptophyta › Embryophyta › Tracheobionta › Magnoliopsida › Apiaceae › Coriandrum › Development and Single Laboratory Evaluation...
questionsmedicales.fr › Infection › Maladies parasitaires › Protozooses › Coccidiose › Cyclosporose › Development and Single Laboratory Evaluation...