Interlaboratory comparison of molecular detection of Cyclospora cayetanensis in routine diagnostic laboratories
10 affiliations
- 1Laboratory for Medical Microbiology and Immunology, Dicoon, Elst, The Netherlands.
- 2Department of Medical Microbiology and Immunology, Rijnstate, Arnhem, The Netherlands.
- 3Laboratory for Medical Microbiology, Medical Laboratory Services, Willemstad, Curaçao.
- 4Department of Medical Microbiology, RadboudUMC, Nijmegen, The Netherlands.
- 5LUCID-M, Leiden University Medical Centre, Leiden, The Netherlands.
- 6Department of Medical Microbiology and Infectious Diseases, Erasmus MC University Medical Center, Rotterdam, The Netherlands.
- 7Microvida, Laboratory for Microbiology and Immunology, Elisabeth-Tweesteden Hospital, Tilburg, The Netherlands.
- 8Laboratory for Medical Microbiology, Unilabs, Amsterdam, The Netherlands.
- 9Department of Medical Microbiology, University Medical Center Utrecht, Utrecht, The Netherlands.
- 10Centre for Infectious Disease Control, National Institute for Public Health and the Environment, Bilthoven, The Netherlands.
Abstract
Objectives: Although molecular detection of Cyclospora cayetanensis is increasingly incorporated into multiplex gastrointestinal diagnostic workflows, dedicated external quality assessment (EQA) programmes for Cyclospora remain limited. We performed a nationwide interlaboratory comparison to evaluate routine molecular detection of C. cayetanensis across decentralized ISO15189-accredited laboratories in the Netherlands. Methods: An anonymized 11-sample panel containing unpreserved stool specimens and eNAT-preserved stool suspensions was distributed to laboratories in the Netherlands (n = 8) and Curaçao (n = 1). Laboratories analysed samples using their routine diagnostic workflows. Workflow characteristics, DNA extraction procedures, molecular platforms, and interpretation criteria were collected by a structured questionnaire. Because detection of Cryptosporidium spp. is usually performed within the same workflow and present similar extraction challenges, Cryptosporidium-positive samples were included as internal workflow controls. Aggregated retrospective laboratory data were collected to explore surveillance feasibility and to gain insight in seasonality of C. cayetanensis and Cryptosporidium spp. Results: Seven laboratories performed molecular detection of C. cayetanensis, while all nine performed detection of Cryptosporidium spp. Complete qualitative agreement was observed for Cryptosporidium spp. detection. For C. cayetanensis, overall qualitative agreement was high, although one false-positive and three false-negative results occurred among core samples. Detection of a low-positive educational sample varied substantially between laboratories, indicating reduced reproducibility near assay-specific limits of detection. This study shows that eNAT-preserved stool suspensions remained stable under ambient transport conditions. Retrospective laboratory data showed low-level year-round C. cayetanensis detection and a seasonal increase in Cryptosporidium spp. positivity during late summer and autumn. Conclusions: Routine molecular detection of C. cayetanensis demonstrated good overall interlaboratory agreement across decentralized diagnostic laboratories. However, reproducibility decreased for low-positive samples, highlighting the importance of assay validation, external quality assessment, and cautious interpretation of weak positive C. cayetanensis PCR results.
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BibTeX
@article{flipse2026interlaboratory,
title = {{Interlaboratory comparison of molecular detection of Cyclospora cayetanensis in routine diagnostic laboratories}},
author = {Jacky Flipse and Angelino T Tromp and Esmée Ruizendaal and Els Wessels and Fleur M H P A Koene and Jaap J van Hellemond and Jaco J Verweij and Linda J Wammes and Noortje van Maarseveen and Raymond Steen and Rob Schuurman and Theun de Groot and Thomas J Roodsant and Wouter Bekers and Guido J H Bastiaens},
journal = {PLOS One},
year = {2026},
month = oct,
doi = {10.1371/journal.pone.0361578},
url = {https://doi.org/10.1371/journal.pone.0361578},
note = {CC BY},
}
RIS
TY - JOUR
TI - Interlaboratory comparison of molecular detection of Cyclospora cayetanensis in routine diagnostic laboratories
AU - Jacky Flipse
AU - Angelino T Tromp
AU - Esmée Ruizendaal
AU - Els Wessels
AU - Fleur M H P A Koene
AU - Jaap J van Hellemond
AU - Jaco J Verweij
AU - Linda J Wammes
AU - Noortje van Maarseveen
AU - Raymond Steen
AU - Rob Schuurman
AU - Theun de Groot
AU - Thomas J Roodsant
AU - Wouter Bekers
AU - Guido J H Bastiaens
PY - 2026
DA - 2026/10/28
JO - PLOS One
DO - 10.1371/journal.pone.0361578
UR - https://doi.org/10.1371/journal.pone.0361578
AB - Objectives: Although molecular detection of Cyclospora cayetanensis is increasingly incorporated into multiplex gastrointestinal diagnostic workflows, dedicated external quality assessment (EQA) programmes for Cyclospora remain limited. We performed a nationwide interlaboratory comparison to evaluate routine molecular detection of C. cayetanensis across decentralized ISO15189-accredited laboratories in the Netherlands. Methods: An anonymized 11-sample panel containing unpreserved stool specimens and eNAT-preserved stool suspensions was distributed to laboratories in the Netherlands (n = 8) and Curaçao (n = 1). Laboratories analysed samples using their routine diagnostic workflows. Workflow characteristics, DNA extraction procedures, molecular platforms, and interpretation criteria were collected by a structured questionnaire. Because detection of Cryptosporidium spp. is usually performed within the same workflow and present similar extraction challenges, Cryptosporidium-positive samples were included as internal workflow controls. Aggregated retrospective laboratory data were collected to explore surveillance feasibility and to gain insight in seasonality of C. cayetanensis and Cryptosporidium spp. Results: Seven laboratories performed molecular detection of C. cayetanensis, while all nine performed detection of Cryptosporidium spp. Complete qualitative agreement was observed for Cryptosporidium spp. detection. For C. cayetanensis, overall qualitative agreement was high, although one false-positive and three false-negative results occurred among core samples. Detection of a low-positive educational sample varied substantially between laboratories, indicating reduced reproducibility near assay-specific limits of detection. This study shows that eNAT-preserved stool suspensions remained stable under ambient transport conditions. Retrospective laboratory data showed low-level year-round C. cayetanensis detection and a seasonal increase in Cryptosporidium spp. positivity during late summer and autumn. Conclusions: Routine molecular detection of C. cayetanensis demonstrated good overall interlaboratory agreement across decentralized diagnostic laboratories. However, reproducibility decreased for low-positive samples, highlighting the importance of assay validation, external quality assessment, and cautious interpretation of weak positive C. cayetanensis PCR results.
SN - 1932-6203
ER -