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Development, Optimisation and Validation of a Novel Multiplex Real-Time PCR Method for the Simultaneous Detection of Cryptosporidium spp., Giardia duodenalis and Dientamoeba fragilis.
Sánchez, Isbene; Dashti, Alejandro; Köster, Pamela C; Bailo, Begoña; González, Nuria; Allende, Janire; Stensvold, Christen Rune; Carmena, David; González-Barrio, David.
Affiliation
  • Sánchez I; Vacunek SL, Bizkaia Technology Park, 48160 Derio, Spain.
  • Dashti A; Parasitology Reference and Research Laboratory, Spanish National Centre for Microbiology, Health Institute Carlos III, Majadahonda, 28220 Madrid, Spain.
  • Köster PC; Parasitology Reference and Research Laboratory, Spanish National Centre for Microbiology, Health Institute Carlos III, Majadahonda, 28220 Madrid, Spain.
  • Bailo B; Parasitology Reference and Research Laboratory, Spanish National Centre for Microbiology, Health Institute Carlos III, Majadahonda, 28220 Madrid, Spain.
  • González N; Vacunek SL, Bizkaia Technology Park, 48160 Derio, Spain.
  • Allende J; Vacunek SL, Bizkaia Technology Park, 48160 Derio, Spain.
  • Stensvold CR; Department of Bacteria, Parasites and Fungi, Infectious Disease Preparedness, Statens Serum Institute, 2300 Copenhagen, Denmark.
  • Carmena D; Parasitology Reference and Research Laboratory, Spanish National Centre for Microbiology, Health Institute Carlos III, Majadahonda, 28220 Madrid, Spain.
  • González-Barrio D; Center for Biomedical Research Network (CIBER) in Infectious Diseases, Health Institute Carlos III, Majadahonda, 28220 Madrid, Spain.
Pathogens ; 11(11)2022 Oct 31.
Article in En | MEDLINE | ID: mdl-36365028
The enteric protozoan parasites Cryptosporidium spp., Giardia duodenalis and Dientamoeba fragilis are-to various extents-contributors to the burden of gastrointestinal illness in high-income countries. Detection of these pathogens by microscopy examination is challenging because of the limited sensitivity and need for specific staining procedures. We developed and optimised a new multiplex real-time PCR assay for the simultaneous detection of Cryptosporidium spp., G. duodenalis and D. fragilis in clinical (stool) samples. The diagnostic performance of the assay was evaluated against a large panel of well-characterised DNA samples positive for Cryptosporidium spp. (n = 126), G. duodenalis (n = 132) and D. fragilis (n = 49). The specificity of the test was assessed against a DNA panel from other intestinal or phylogenetically related parasites (n = 105) and faecal DNA from individuals without clinical manifestations (n = 12). The assay exhibited a diagnostic sensitivity of 0.90-0.97 and a diagnostic specificity of 1. The limit of detection was estimated for Cryptosporidium (1 oocyst) and G. duodenalis (5 × 10-4 cysts). The method allowed the detection of four Cryptosporidium species (C. hominis, C. parvum, C. meleagridis and C. cuniculus) and five G. duodenalis assemblages (A-E) without cross-reacting with other parasites belonging to the phyla Amoebozoa, Apicomplexa, Euglenozoa, Microsporidia, Nematoda and Platyhelminthes. This newly developed multiplex real-time PCR assay represents a novel alternative for the rapid and accurate detection of Cryptosporidium, G. duodenalis and D. fragilis in clinical settings.
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Full text: 1 Collection: 01-internacional Database: MEDLINE Type of study: Diagnostic_studies Language: En Journal: Pathogens Year: 2022 Document type: Article Affiliation country: Country of publication:

Full text: 1 Collection: 01-internacional Database: MEDLINE Type of study: Diagnostic_studies Language: En Journal: Pathogens Year: 2022 Document type: Article Affiliation country: Country of publication: