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  • Title: Evaluation of a Novel Commercial Real-Time PCR Assay for the Simultaneous Detection of Cryptosporidium spp., Giardia duodenalis, and Entamoeba histolytica.
    Author: Dashti A, Alonso H, Escolar-Miñana C, Köster PC, Bailo B, Carmena D, González-Barrio D.
    Journal: Microbiol Spectr; 2022 Jun 29; 10(3):e0053122. PubMed ID: 35502919.
    Abstract:
    Cryptosporidium spp., Giardia duodenalis, and Entamoeba histolytica are the most common diarrhea-causing protozoan species globally. Misdiagnosis is a concern for asymptomatic and chronic infections. Multiplexing, i.e., the detection of more than one parasite in a single test by real-time PCR, allows high diagnostic performance with favorable cost-effectiveness. We conducted a clinical evaluation of the VIASURE Cryptosporidium, Giardia, & E. histolytica real-time PCR assay (CerTest Biotec, San Mateo de Gállego, Spain) against a large panel (n = 358) of well-characterized DNA samples positive for Cryptosporidium spp. (n = 96), G. duodenalis (n = 115), E. histolytica (n = 25), and other parasitic species of the phyla Amoebozoa (n = 11), Apicomplexa (n = 14), Euglenozoa (n = 8), Heterokonta (n = 42), Metamonada (n = 37), Microsporidia (n = 4), and Nematoda (n = 6). DNA samples were obtained from clinical stool specimens or cultured isolates in a national reference center. Estimated sensitivity and specificity were 0.96 and 0.99 for Cryptosporidium spp., 0.94 and 1 for G. duodenalis, and 0.96 and 1 for E. histolytica, respectively. Positive and negative predictive values were calculated as 1 and 0.98 for Cryptosporidium spp., 0.99 and 0.98 for G. duodenalis, and 1 and 0.99 for E. histolytica, respectively. The assay identified six Cryptosporidium species (Cryptosporidium hominis, Cryptosporidium parvum, Cryptosporidium canis, Cryptosporidium felis, Cryptosporidium scrofarum, and Cryptosporidium ryanae) and four G. duodenalis assemblages (A, B, C, and F). The VIASURE assay provides rapid and accurate simultaneous detection and identification of the most commonly occurring species and genetic variants of diarrhea-causing parasitic protozoa in humans. IMPORTANCE Thorough independent assessment of the diagnostic performance of novel diagnostic assays is essential to ascertain their true usefulness and applicability in routine clinical practice. This is particularly true for commercially available kits based on multiplex real-time PCR aimed to detect and differentiate multiple pathogens in a single biological sample. In this study, we conducted a clinical evaluation of the VIASURE Cryptosporidium, Giardia, & E. histolytica real-time PCR assay (CerTest Biotec) for the detection and identification of the diarrhea-causing enteric protozoan parasites Cryptosporidium spp., G. duodenalis, and E. histolytica. A large panel of well-characterized DNA samples from clinical stool specimens or cultured isolates from a reference center was used for this purpose. The VIASURE assay demonstrated good performance for the routine testing of these pathogens in clinical microbiological laboratories.
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