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1. Fumagillin Shortage: How to Treat Enterocytozoon bieneusi Microsporidiosis in Solid Organ Transplant Recipients in 2024?

Author

Cyril Garrouste, Philippe Poirier, Charlotte Uro-Coste, Xavier Iriart, Nassim Kamar, Julie Bonhomme, Eve Calvar, Solène Le Gal, Luca Lanfranco, Brice Autier, Lucien Rakoff, Marie-Fleur Durieux, Clément Danthu, Florent Morio, Clément Deltombe, Alicia Moreno-Sabater, Nacera Ouali, Damien Costa, Dominique Bertrand, Adélaïde Chesnay, Philippe Gatault, Meja Rabodonirina, Emmanuel Morelon, Jérôme Dumortier, Emilie Sitterlé, Anne Scemla, Samia Hamane, Laurène Cachera, Céline Damiani, Coralie Poulain, Coralie L’Ollivier, Valérie Moal, Laurence Delhaes, Hannah Kaminski, Estelle Cateau, Laure Ecotière, Julie Brunet, Sophie Caillard, Stéphane Valot, Claire Tinel, Nicolas Argy, Quentin Raimbourg, Marie Gladys Robert, Johan Noble, Aude Boignard, Françoise Botterel, Marie Matignon, Anne-Pauline Bellanger, Thomas Crépin, Jordan Leroy, Arnaud Lionet, Anne Debourgogne, Muriel Nicolas, Joëlle Claudéon, Maxime Moniot, Céline Lambert, and Céline Nourrisson

Subjects
solid organ transplant, nitazoxanide, microsporidiosis, Enterocytozoon bieneusi, infectious diarrhea, Specialties of internal medicine, RC581-951
Abstract

Intestinal microsporidiosis caused by Enterocytozoon bieneusi is an opportunistic infection that especially affects solid organ transplant (SOT) recipients. Management revolves around tapering the immunosuppressive regimen and/or using a specific anti-microsporidia treatment, but only fumagillin has demonstrated efficacy for treatment of this infection. Since fumagillin has been commercially discontinued, nitazoxanide is increasingly being used in this indication. We aimed to describe therapeutic management of E. bieneusi infections in this context. We conducted a French nationwide observational retrospective study on reported cases of E. bieneusi infections in SOT recipients. We identified 154 cases: 64 (41.6%) were managed by simply modifying the immunosuppressive regimen, 54 (35.1%) were given fumagillin, and 36 (23.4%) were given nitazoxanide. Clinical remission rate ranged from 77.8% to 90.7% and was not significantly different between therapeutic strategies but tended to be lower with nitazoxanide. Stool negativization rate was highest with fumagillin (91.7%) and lowest with nitazoxanide (28.6%). Relapses occurred in 6.9% of cases and were more frequent with nitazoxanide (14.3%). This study shows that tapering immunosuppression can result in a satisfactory remission rate but is sometimes accompanied by relapses. Nitazoxanide had limited effectiveness, whereas fumagillin had good results that provide a solid rationale for bringing fumagillin back to market.Trial Registration NumberClinicalTrials.gov ID: NCT05417815.

Published
2024
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2. Summer diarrhea in children: a monocentric French epidemiological observational study

Author

Camille Mallier, Elisa Creuzet, Céline Lambert, Julien Delmas, Audrey Mirand, Emmanuelle Rochette, Stéphane Valot, Maxime Moniot, Frédéric Dalle, Cécile Henquell, Etienne Merlin, Philippe Poirier, Matthieu Verdan, and Céline Nourrisson

Subjects
Medicine, Science
Abstract

Abstract Pediatric diarrhea is a major public health problem worldwide. In France, continuous surveillance shows a winter epidemic peak and a more modest summer recrudescence. Few studies describe the infectious agents responsible for pediatric summer diarrhea in France. The objectives were to estimate the prevalence of infectious diarrhea and describe the pathogens responsible for summer diarrhea in children; and to describe common factors that can be used as guidance on the etiology of these diarrheas. A cross-sectional, single-center, epidemiological observational study was conducted in the pediatric emergency department of a French hospital between June and September in 2019 and 2020. Multiplex gastrointestinal pathogen panels were used for diagnostics. A multiple correspondence analysis was used to determine profiles of patients. A total of 95 children were included, of whom 82.1% (78/95) were under five years old. The prevalence of infectious summer diarrhea was 81.1% (77/95, 95%CI 71.7–88.4%). A total of 126 infectious agents were detected (50.0% bacteria, 38.1% viruses, 11.9% parasites). The main enteric pathogens were enteropathogen Escherichia coli (24/126), rotavirus (17/126) and Salmonella (16/126). A co-detection was found in 51.9% (40/77) of cases. Four patient profiles, considering the severity and the pathogen involved, were highlighted.

Published
2023
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4. A summary of cryptosporidiosis outbreaks reported in France and overseas departments, 2017–2020

Author

Damien Costa, Romy Razakandrainibe, Louise Basmaciyan, Jérôme Raibaut, Pascal Delaunay, Florent Morio, Gilles Gargala, Venceslas Villier, Abdelmounaim Mouhajir, Bernard Levy, Catherine Rieder, Sébastien Larreche, Sophie Lesthelle, Noémie Coron, Estelle Menu, Magalie Demar, Vincent Pommier de Santi, Véronique Blanc, Stéphane Valot, Frédéric Dalle, and Loic Favennec

Subjects
Cryptosporidium, Outbreak, France, Waterborne, Foodborne, Infectious and parasitic diseases, RC109-216
Abstract

Cryptosporidium is a known foodborne pathogen, ranked fifth out of 24 among foodborne parasites in terms of importance and a cause of many cryptosporidiosis outbreaks worldwide. In France, very few outbreaks were reported before 2017, and data recently obtained by the Expert Laboratory of the Cryptosporidiosis National Reference Center (CNR-LE-Cryptosporidiosis) have shown that outbreaks are in fact common and frequently underreported. In this work, we aim to report the characteristics of outbreaks detected in France during the period 2017–2020 and present a summary of investigations carried out by the CNR-LE-Cryptosporidiosis. During the study period, there were eleven cryptosporidiosis outbreaks, including three with no identified origin. Among the eight identified outbreaks: six were due to water contamination (five tap water and one recreational water), one was due to direct contact with infected calves, and one was due to consumption of contaminated curd cheese. Among these outbreaks, five of them exceeded one hundred cases. Recent results obtained by the CNR-LE-Cryptosporidiosis revealed the multiannual occurrence of Cryptosporidium outbreaks in France. Waterborne outbreaks were more frequently detected, while foodborne outbreaks which are more difficult to detect were likely underreported.

Published
2022
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5. Cryptosporidiosis outbreak in Amazonia, French Guiana, 2018

Author

Estelle Menu, Emilie Mosnier, Arnaud Cotrel, Loic Favennec, Romy Razakandrainibe, Stéphane Valot, Denis Blanchet, Frédéric Dalle, Damien Costa, Mélanie Gaillet, Magalie Demar, and Franck de Laval

Subjects
Arctic medicine. Tropical medicine, RC955-962, Public aspects of medicine, RA1-1270
Abstract

Background Cryptosporidiosis outbreaks in South America are poorly documented. In March 2018, 51 cases of cryptosporidiosis were reported in Maripasoula, a village located in a remote forest area along the border between Surinam and French Guiana. Method To identify the origin of the epidemic, we performed epidemiological, microbiological, and environmental investigations. Only the cases involving diarrhoea and Cryptosporidium-positive stool were considered as bona fide, while cases involving diarrhoea and close contact with a confirmed case were classified as “possible”. Results We identified 16 confirmed cases and 35 possible ones. Confirmed cases comprised nine children (median age of 18 months, range: 6–21), one immunocompromised adult and six soldiers. One child required a hospitalisation for rehydration. All 16 Cryptosporidium stools were PCR positive, and sequencing of the gp60 gene confirmed only one Cryptosporidium hominis subtype IbA10G2. Tap water consumption was the only common risk factor identified. Contamination of the water network with Cryptosporidium parvum subtype IIdA19G2 was found. Conclusion Water quality is a major public health issue in Amazonian French Guiana, especially for population at risk (children, people with comorbidity, travelers). For them, alternative water supply or treatment should be implemented.

Published
2022

6. Evaluation of Next-Generation Sequencing Applied to Cryptosporidium parvum and Cryptosporidium hominis Epidemiological Study

Author

Eloïse Bailly, Stéphane Valot, Anne Vincent, Yannis Duffourd, Nadège Grangier, Martin Chevarin, Damien Costa, Romy Razakandrainibe, Loïc Favennec, Louise Basmaciyan, and Frédéric Dalle

Subjects
Next-Generation Sequencing, Cryptosporidium sp., cryptosporidiosis, epidemiology, subtyping, genetic diversity, Medicine
Abstract

Background. Nowadays, most of the C. parvum and C. hominis epidemiological studies are based on gp60 gene subtyping using the Sanger sequencing (SgS) method. Unfortunately, SgS presents the limitation of being unable to detect mixed infections. Next-Generation Sequencing (NGS) seems to be an interesting solution to overcome SgS limits. Thus, the aim of our study was to (i) evaluate the reliability of NGS as a molecular typing tool for cryptosporidiosis, (ii) investigate the genetic diversity of the parasite and the frequency of mixed infections, (iii) assess NGS usefulness in Cryptosporidium sp. outbreak investigations, and (iv) assess an interpretation threshold of sequencing data. Methods. 108 DNA extracts from positive samples were sequenced by NGS. Among them, two samples were used to validate the reliability of the subtyping obtained by NGS and its capacity to detect DNA mixtures. In parallel, 106 samples from French outbreaks were used to expose NGS to epidemic samples. Results. NGS proved suitable for Cryptosporidium sp. subtyping at the gp60 gene locus, bringing more genetic information compared to SgS, especially by working on many samples simultaneously and detecting more diversity. Conclusions. This study confirms the usefulness of NGS applied to C. hominis and C. parvum epidemiological studies, especially aimed at detecting minority variants.

Published
2022
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7. Disseminated Cryptosporidium infection in an infant with CD40L deficiency

Author

Fleur Dupuy, Stéphane Valot, Frédéric Dalle, Arthur Sterin, and Coralie L’Ollivier

Subjects
Cryptosporidium, Disseminated, CD40L deficiency, Infectious and parasitic diseases, RC109-216
Abstract

The protozoan Cryptosporidium affects the digestive tract of humans and animals. Cryptosporidiosis leads to diarrhoea mimicking a cholera-like course with dehydration and may even result in death in immunodeficient patients, as patients with hyper-IgM syndrome. We describe a rare case of disseminated Cryptosporidium infection in a seven- year-old boy with CD40 L deficiency. During the pre-graft phase, the patient presented an intestinal cryptosporidiosis which became complicated few days later during the aplasia period with a cholangitis and pulmonary cryptosporidiosis. Cryptosporidium hominis was identified. After treatment with nitazoxanide and azithromycine the patient was doing well.

Published
2021
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8. Oestrus ovis external ophtalmomyiasis: a case report in Burgundy France

Author

Louise Basmaciyan, Pierre-Henry Gabrielle, Stéphane Valot, Marc Sautour, Jean-Christophe Buisson, Catherine Creuzot-Garcher, and Frédéric Dalle

Subjects
Ophtalmomyiasis, Oestrus ovis, Burgundy, Ophthalmology, RE1-994
Abstract

Abstract Background External ophtalmomyiasis (EOM) is a zoonosis related to the presence of Oestrus ovis larvae at the ocular level in small ruminants (i.e. ovine, caprine). In humans, EOM is a rare cosmopolitan disorder, mostly described in warm and dry rural areas in patients living close to livestock areas. In metropolitan France (excluding Corsica), EOM is an exceptional disease with less than 25 cases recorded since 1917. Case presentation We report a case of EOM in a 19-years old man in the last week of September 2016 in Burgundy. Conclusion The diagnosis of an EOM in Burgundy, a French region described as cold and humid, is surprising and could be due to a more marked climatic warming during the vegetative season in Burgundy resulting in the implantation of Diptera of the genus Oestrus sp. in this region.

Published
2018
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9. Commercial Simplex and Multiplex PCR Assays for the Detection of Intestinal Parasites Giardia intestinalis, Entamoeba spp., and Cryptosporidium spp.: Comparative Evaluation of Seven Commercial PCR Kits with Routine In-House Simplex PCR Assays

Author

Louise Basmaciyan, Alexandre François, Anne Vincent, Stéphane Valot, Alain Bonnin, Damien Costa, Romy Razakandrainibe, Florent Morio, Loic Favennec, and Frédéric Dalle

Subjects
intestinal parasitic diseases, Entamoeba spp., Giardia intestinalis, Cryptosporidium spp., diagnosis, DNA amplification, Biology (General), QH301-705.5
Abstract

Nowadays, many commercial kits allowing the detection of digestive parasites by DNA amplification methods have been developed, including simplex PCR assays (SimpPCRa) allowing the identification of a single parasite, and multiplex PCR assays (MultPCRa) allowing the identification of several parasites at once. Thus, aimed at improving the diagnosis of intestinal protozoal infections, it is essential to evaluate the performances of these new tools. A total of 174 DNA samples collected between 2007 and 2017 were retrospectively included in this study. Performances of four commercial SimpPCRa (i.e., CerTest-VIASURETM) and three MultPCRa (i.e., CerTest-VIASURETM, FAST-TRACK-Diagnostics-FTD-Stool-ParasiteTM and DIAGENODE-Gastroenteritis/Parasite-panel-ITM) were evaluated for the detection of Cryptosporidium spp., Entamoeba spp., and Giardia intestinalis in stool samples compared to our routinely used in-house SimpPCRa. Globally, the SimpPCRa showed better sensitivity/specificity for the detection of G. intestinalis, E. histolytica, E. dispar, and Cryptosporidium spp. (i.e., 96.9/93.6%; 100/100%; 95.5/100%; and 100/99.3%, respectively), compared to the three commercial MultPCRa tested. All in all, we showed that MultPCRa offer an interesting alternative for the detection of protozoans in stool samples depending on the clinical context.

Published
2021
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10. Comparative Performance of Eight PCR Methods to Detect Cryptosporidium Species

Author

Damien Costa, Louise Soulieux, Romy Razakandrainibe, Louise Basmaciyan, Gilles Gargala, Stéphane Valot, Frédéric Dalle, and Loic Favennec

Subjects
Cryptosporidium, PCR, detection, diagnosis, sensitivity, specificity, Medicine
Abstract

Diagnostic approaches based on PCR methods are increasingly used in the field of parasitology, particularly to detect Cryptosporidium. Consequently, many different PCR methods are available, both “in-house” and commercial methods. The aim of this study was to compare the performance of eight PCR methods, four “in-house” and four commercial methods, to detect Cryptosporidium species. On the same DNA extracts, performance was evaluated regarding the limit of detection for both C. parvum and C. hominis specificity and the ability to detect rare species implicated in human infection. Results showed variations in terms of performance. The best performance was observed with the FTD® Stool parasites method, which detected C. parvum and C. hominis with a limit of detection of 1 and 10 oocysts/gram of stool respectively; all rare species tested were detected (C. cuniculus, C. meleagridis, C. felis, C. chipmunk, and C. ubiquitum), and no cross-reaction was observed. In addition, no cross-reactivity was observed with other enteric pathogens. However, commercial methods were unable to differentiate Cryptosporidium species, and generally, we recommend testing each DNA extract in at least triplicate to optimize the limit of detection.

Published
2021
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11. Comparative Study of Eleven Mechanical Pretreatment Protocols for Cryptosporidium parvum DNA Extraction from Stool Samples

Author

Laure Claudel, Nicolas Valeix, Louise Basmaciyan, Bruno Pereira, Damien Costa, Anne Vincent, Stéphane Valot, Loic Favennec, and Frederic Dalle

Subjects
Cryptosporidium parvum, mechanical pretreatment, DNA extraction, stool samples, real-time PCR, molecular diagnosis, Biology (General), QH301-705.5
Abstract

Nowadays, many commercial kits allow the polymerase chain reaction (PCR) detection of Cryptosporidium deoxyribonucleic acid (DNA) in stool samples, the efficiency of which relies on the extraction method used. Mechanical pretreatment of the stools using grinding beads has been reported to greatly improve this extraction step. However, optimization of this key step remains to be carried out. Indeed, many parameters could influence the pretreatment performances, among which the modulation of the speed and duration of the grinding step, in addition to the physicochemical features of the grinding beads, have never been evaluated to date. In this study, eleven commercial mechanical pretreatment matrixes (Lysis matrix tubes®, MP Biomedical, Irvine, CA, USA) composed of beads with different sizes, shapes, and molecular compositions, were evaluated for their performances in improving Cryptosporidium parvum oocyst DNA extraction before amplification by using our routinely used real-time PCR method. As expected, the eleven commercial mechanical pretreatment matrixes showed varying performances depending on the composition, size, and shape. All in all, the best performances were obtained when using the Lysing matrix, including ceramic beads with a median size (diameter of 1.4 mm).

Published
2021
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12. Epidemiology of Cryptosporidiosis in France from 2017 to 2019

Author

Damien Costa, Romy Razakandrainibe, Stéphane Valot, Margot Vannier, Marc Sautour, Louise Basmaciyan, Gilles Gargala, Venceslas Viller, Denis Lemeteil, Jean-Jacques Ballet, French National Network on Surveillance of Human Cryptosporidiosis, Frédéric Dalle, and Loïc Favennec

Subjects
cryptosporidiosis, epidemiology, France, Biology (General), QH301-705.5
Abstract

Cryptosporidiosis is currently recognized worldwide as a leading cause of moderate to severe diarrhea. In Europe, large water- and foodborne outbreaks have been reported, highlighting the widespread distribution of the parasite and its important health impact. Surveillance networks have been progressively set up and the aim of this study was to present recent epidemiological data obtained in France from 2017 to 2019 by the National Reference Center—Expert Laboratory of cryptosporidiosis (Centre National de Référence–Laboratoire Expert cryptosporidioses CNR-LE). Data were obtained from online reports of volunteer network participants and stools were sent to the CNR-LE for species identification and GP60 genotyping. During this period, data from 750 online reports were available. Cryptosporidiosis occurred predominantly in young children (Cryptosporidium parvum was largely predominant (72% of cases) over C. hominis (24%) and some other uncommon species. C. parvum GP60 subtypes IIa and IId were the most represented, which suggests frequent zoonotic transmission. For C. hominis, subtypes IbA10G2 and IaA22R2 were predominant.

Published
2020
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13. Multicenter Comparative Study of Six Cryptosporidium parvum DNA Extraction Protocols Including Mechanical Pretreatment from Stool Samples

Author

Nicolas Valeix, Damien Costa, Louise Basmaciyan, Stéphane Valot, Anne Vincent, Romy Razakandrainibe, Florence Robert-Gangneux, Céline Nourrisson, Bruno Pereira, Emilie Fréalle, Philippe Poirier, Loic Favennec, and Frederic Dalle

Subjects
Cryptosporidium parvum, grinding, DNA extraction, stool samples, real-time PCR, molecular diagnosis, Biology (General), QH301-705.5
Abstract

Background: Nowadays, many commercial kits allow the detection of Cryptosporidium sp. in stool samples after deoxyribonucleic acid (DNA) extraction. Protocols of stool pretreatment have been proposed to optimize oocysts’ DNA extraction. Among them, mechanical grinding was reported to improve the performance of Cryptosporidium oocysts’ DNA extraction. Methods: A multicenter comparative study was conducted within the framework of the French National Reference Center-Expert Laboratory for Cryptosporidiosis. Six extraction systems (i.e., manual or automated) associated with various mechanical pretreatment protocols, were compared for the Cryptosporidium parvum oocyst’ DNA extraction, before amplification using the same real-time PCR method targeting. Results: The sensitivity of real-time PCR assay was unequally impacted by the pretreatment/extraction protocol. We observed significant differences for the lowest concentrations of C. parvum oocysts (i.e., 0–94.4% and 33.3–100% respectively for 10 and 50 oocysts/mL). All in all, the protocol using Quick DNA Fecal/Soil Microbe-Miniprep® manual kit showed the best performances. In addition, optimal performances of mechanical pretreatment were obtained by combining a grinding duration of 60 s with a speed of 4 m/s using Fastprep24® with Lysing Matrix E®. Conclusions: Sample pretreatment, as well as the extraction method, needs to be properly adapted to improve the diagnostic performances of the C. parvum DNA amplification methods.

Published
2020
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14. Diagnosis of mucormycosis using an intercalating dye-based quantitative PCR

Author

Jeanne Bigot, Alexandre Godmer, Lysa Prudenté, Cécile Angebault, Eolia Brissot, Naike Bige, Guillaume Voiriot, Pierre-Louis Leger, Camille Petit-Hoang, Sarah Atallah, Elodie Gouache, Yaye Senghor, Stéphane Valot, Christophe Hennequin, and Juliette Guitard

Subjects
Infectious Diseases, Mucorales, Animals, Mucormycosis, General Medicine, DNA, Fungal, Real-Time Polymerase Chain Reaction, DNA Primers
Abstract

PCR-based methods applied to various body fluids emerged in recent years as a promising approach for the diagnosis of mucormycosis. In this study, we set up and assess the value of a qPCR to detect a wide variety of Mucorales species in a single tube. A pair of degenerated primers targeting the rDNA operon was used in a qPCR utilizing an intercalating fluorescent dye. Analytical assessment, using a wide variety of both Mucorales strains (8 genera, 11 species) and non-Mucorales strains (9 genera, 14 species), showed 100% sensitivity and specificity rates with a limit of detection at 3 rDNA copy/qPCR reaction. Subsequently, 364 clinical specimens from 166 at-risk patients were prospectively tested with the assay. All the seven patients classified as proven/probable mucormycosis using the EORTC-MSG criteria had a positive qPCR as well as a patient with a proven uncharacterized invasive mold infection. In addition, three out of seven patients with possible mold invasive infections had at least one positive qPCR test. Sensitivity was calculated between 73.33 and 100% and specificity between 98.10 and 100%. The qPCR method proposed showed excellent performances and would be an important adjunctive tool for the difficult diagnosis of mucormycosis diagnosis. Lay abstract qPCR-based diagnosis is the most reliable approach for mucormycosis. We set up a pan-Mucorales qPCR able to detect in a single reaction not less than 11 different species. Both analytical and clinical performances support its use in the clinical setting.

Published
2022
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15. Commercial Simplex and Multiplex PCR Assays for the Detection of Intestinal Parasites Giardia intestinalis, Entamoeba spp., and Cryptosporidium spp.: Comparative Evaluation of Seven Commercial PCR Kits with Routine In-House Simplex PCR Assays

Author

Damien Costa, Alexandre François, Anne Vincent, Romy Razakandrainibe, Alain Bonnin, Florent Morio, Stéphane Valot, Frédéric Dalle, Loïc Favennec, and Louise Basmaciyan

Subjects
Microbiology (medical), diagnosis, QH301-705.5, Dispar, Pcr assay, Context (language use), Biology, Giardia intestinalis, Cryptosporidium spp, Microbiology, stool samples, Article, Entamoeba spp, DNA amplification, Virology, Multiplex polymerase chain reaction, parasitic diseases, Parasite hosting, Biology (General), Entamoeba, Cryptosporidium, biology.organism_classification, PCR, intestinal parasitic diseases
Abstract

Nowadays, many commercial kits allowing the detection of digestive parasites by DNA amplification methods have been developed, including simplex PCR assays (SimpPCRa) allowing the identification of a single parasite, and multiplex PCR assays (MultPCRa) allowing the identification of several parasites at once. Thus, aimed at improving the diagnosis of intestinal protozoal infections, it is essential to evaluate the performances of these new tools. A total of 174 DNA samples collected between 2007 and 2017 were retrospectively included in this study. Performances of four commercial SimpPCRa (i.e., CerTest-VIASURETM) and three MultPCRa (i.e., CerTest-VIASURETM, FAST-TRACK-Diagnostics-FTD-Stool-ParasiteTM and DIAGENODE-Gastroenteritis/Parasite-panel-ITM) were evaluated for the detection of Cryptosporidium spp., Entamoeba spp., and Giardia intestinalis in stool samples compared to our routinely used in-house SimpPCRa. Globally, the SimpPCRa showed better sensitivity/specificity for the detection of G. intestinalis, E. histolytica, E. dispar, and Cryptosporidium spp. (i.e., 96.9/93.6%, 100/100%, 95.5/100%, and 100/99.3%, respectively), compared to the three commercial MultPCRa tested. All in all, we showed that MultPCRa offer an interesting alternative for the detection of protozoans in stool samples depending on the clinical context.

Published
2021

16. Selecting a multiplex PCR panel for accurate molecular diagnosis of intestinal protists: a comparative study of Allplex

Author

Nicolas, Argy, Céline, Nourrisson, Ahmed, Aboubacar, Philippe, Poirier, Stéphane, Valot, Adrien, Laude, Guillaume, Desoubeaux, Christelle, Pomares, Marie, Machouart, Yohann, Le Govic, Frédéric, Dalle, Françoise, Botterel, Nathalie, Bourgeois, Estelle, Cateau, Marion, Leterrier, Patrice, Le Pape, Florent, Morio, and Sandrine, Houze

Subjects
Feces, Sheep, Entamoeba histolytica, Animals, Cryptosporidiosis, Cryptosporidium, Giardia lamblia, Multiplex Polymerase Chain Reaction, Sensitivity and Specificity, Retrospective Studies, Scrapie
Abstract

Commercial multiplex PCR assay panels were developed to overcome the limitations of microscopic examination for parasitological diagnosis on stool samples. However, given the increased supply of this diagnostic approach, these assays must be evaluated to position them in a diagnostic algorithm. Analytical performances of the multiplex PCR assay G-DiaParaTrio, AllplexSélection d’un panel PCR multiplex pour un diagnostic moléculaire précis des protistes intestinaux : étude comparative des tests AllplexDes panels commerciaux de tests PCR multiplex ont été développés pour dépasser les limites de l’examen microscopique pour l’examen parasitologique des selles. Cependant, compte tenu de l’offre croissante de cette approche diagnostique, ces tests doivent être évalués pour les positionner dans un algorithme de diagnostic. Les performances analytiques des tests PCR multiplex G-DiaParaTrio, Allplex

Published
2021

17. Comparative Study of Eleven Mechanical Pretreatment Protocols for

Author

Laure, Claudel, Nicolas, Valeix, Louise, Basmaciyan, Bruno, Pereira, Damien, Costa, Anne, Vincent, Stéphane, Valot, Loic, Favennec, and Frederic, Dalle

Subjects
Cryptosporidium parvum, molecular diagnosis, mechanical pretreatment, real-time PCR, DNA extraction, stool samples, Article
Abstract

Nowadays, many commercial kits allow the polymerase chain reaction (PCR) detection of Cryptosporidium deoxyribonucleic acid (DNA) in stool samples, the efficiency of which relies on the extraction method used. Mechanical pretreatment of the stools using grinding beads has been reported to greatly improve this extraction step. However, optimization of this key step remains to be carried out. Indeed, many parameters could influence the pretreatment performances, among which the modulation of the speed and duration of the grinding step, in addition to the physicochemical features of the grinding beads, have never been evaluated to date. In this study, eleven commercial mechanical pretreatment matrixes (Lysis matrix tubes®, MP Biomedical, Irvine, CA, USA) composed of beads with different sizes, shapes, and molecular compositions, were evaluated for their performances in improving Cryptosporidium parvum oocyst DNA extraction before amplification by using our routinely used real-time PCR method. As expected, the eleven commercial mechanical pretreatment matrixes showed varying performances depending on the composition, size, and shape. All in all, the best performances were obtained when using the Lysing matrix, including ceramic beads with a median size (diameter of 1.4 mm).

Published
2020

18. Actualités du diagnostic des infections parasitaires et fongiques

Author

Florent Morio, Arezki Izri, Emilie Fréalle, Renaud Piarroux, Laurence Lachaud, Odile Villard, Stéphane Valot, Eric Dannaoui, Hélène Yera, Jean Menotti, Florence Persat, Sandrine Houzé, CHU Lille, Centre d’Infection et d’Immunité de Lille - INSERM U 1019 - UMR 9017 - UMR 8204 (CIIL), Institut Pasteur de Lille, Réseau International des Instituts Pasteur (RIIP)-Réseau International des Instituts Pasteur (RIIP)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Université de Lille-Centre Hospitalier Régional Universitaire [Lille] (CHRU Lille)-Centre National de la Recherche Scientifique (CNRS), Laboratoire de parasitologie mycologie (CHU de Dijon), Centre Hospitalier Universitaire de Dijon - Hôpital François Mitterrand (CHU Dijon), Institut Pierre Louis d'Epidémiologie et de Santé Publique (iPLESP), Institut National de la Santé et de la Recherche Médicale (INSERM)-Sorbonne Université (SU), Epidémiologie, Systèmes d'Information, Modélisation, Université Pierre et Marie Curie - Paris 6 (UPMC)-Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Institut National de la Santé et de la Recherche Médicale (INSERM)-CHU Pitié-Salpêtrière [AP-HP], Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Sorbonne Université (SU)-Sorbonne Université (SU), Institut des Agents Infectieux [Lyon] (IAI), Hospices Civils de Lyon (HCL), Laboratoire de Parasitologie-Mycologie (CHU de Montpellier), Pôle Biologie-Pathologie [CHRU Montpellier], Centre Hospitalier Régional Universitaire [Montpellier] (CHRU Montpellier)-Centre Hospitalier Régional Universitaire [Montpellier] (CHRU Montpellier), Maladies infectieuses et vecteurs : écologie, génétique, évolution et contrôle (MIVEGEC), Université de Montpellier (UM)-Centre National de la Recherche Scientifique (CNRS)-Institut de Recherche pour le Développement (IRD [France-Sud]), Service de Parasitologie [Avicenne], Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Hôpital Avicenne [AP-HP], Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Université Sorbonne Paris Nord, Laboratoire de Parasitologie et de Mycologie Médicale [Strasbourg], Les Hôpitaux Universitaires de Strasbourg (HUS), Service de parasitologie-mycologie [CHU Cochin], Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Hôpital Cochin [AP-HP], Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP), Hôpital Européen Georges Pompidou [APHP] (HEGP), Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Hôpitaux Universitaires Paris Ouest - Hôpitaux Universitaires Île de France Ouest (HUPO), Centre hospitalier universitaire de Nantes (CHU Nantes), Cibles et Médicaments des Infections et de l'Immunité (IICiMed), Université de Nantes - UFR des Sciences Pharmaceutiques et Biologiques, Université de Nantes (UN)-Université de Nantes (UN), AP-HP - Hôpital Bichat - Claude Bernard [Paris], Mère et enfant en milieu tropical : pathogènes, système de santé et transition épidémiologique (MERIT - UMR_D 261), and Institut de Recherche pour le Développement (IRD)-Université Paris Cité (UPCité)

Subjects
medicine.medical_specialty, Molecular biology, MESH: Parasitology, MESH: Mycology, Mycology, MESH: Parasitic Diseases, Serology, Fungal infections, MESH: Mycoses, MESH: Laboratories, Hospital, medicine, Humans, Medical physics, Diagnostic Techniques and Procedures, MESH: Diagnostic Techniques and Procedures, MESH: Humans, Mass spectrometry, business.industry, MESH: Clinical Laboratory Services, General Medicine, Clinical Laboratory Services, Laboratories, Hospital, Parasitic diseases, [SDV.MP]Life Sciences [q-bio]/Microbiology and Parasitology, Mycoses, Parasitology, business
Abstract

International audience; The diagnosis of parasitic and fungal infections, historically based on the detection of these pathogens using direct diagnosis (macro/microscopic examination, culture) or serological methods, has considerably evolved in the last decades, especially with the development of molecular approaches and mass spectrometry. These techniques, as well as most analyses of parasitic and fungal serology, are mostly the preserve of Hospital University Centers Parasitology-Mycology laboratories. In 2016, the French association of medical parasitology and mycology teachers and hospital practitioners (Anofel) has provided a Catalogue of rare analyses, regularly updated and freely accessible on the Anofel website (https://anofel.net/). This tool, which hinges on 4 parts (parasitology, parasitic serology, mycology, and fungal serology), aims to provide information on all available analyses, and a list of hospital laboratories able to undertake them. It is complementary to the other reference works that were developed by our association, including the Guide of analyses and methods in parasitology and mycology, published in 2018, and the eANOFEL pictures and videos database, freely accessible online (http://www.eanofel.fr). In this article, we draw-up a state-of-the-art of the most specialized techniques available in the parasitology-mycology laboratories and presented in the Catalogue of rare analyses of the Anofel collegium, and their interest for the diagnosis of these infections.; Le diagnostic des infections parasitaires et fongiques, historiquement basé sur la détection de ces pathogènes par diagnostic direct (examen macro/microscopique, culture) ou sérologique, a connu durant les dernières décennies des évolutions importantes, avec notamment le développement d’approches moléculaires et de la spectrométrie de masse. Ces techniques, de même que la plupart des analyses en sérologie parasitaire et fongique, restent le plus souvent l’apanage des laboratoires de centres hospitaliers universitaires spécialisés en parasitologie-mycologie. En 2016, l’Association française des enseignants et praticiens hospitaliers titulaires de parasitologie et mycologie médicales (Anofel) a mis à disposition un Catalogue des analyses rares en accès libre sur le site d’Anofel (https://anofel.net/), et régulièrement mis à jour. Cet outil, qui s’articule en 4 volets (parasitologie, sérologie parasitaire, mycologie et sérologie fongique) a pour objectif de faciliter l’accès aux informations sur les analyses disponibles et les laboratoires hospitaliers capables de les prendre en charge. Il complète les autres ouvrages de référence élaborés par la collégiale Anofel, dont le Guide des analyses et méthodes en parasitologie et mycologie, publié en 2018, et la base de données d’images et de vidéos, en accès libre sur internet (http://www.eanofel.fr). L’objectif de cet article est de dresser un état des lieux des techniques plus spécialisées disponibles dans les laboratoires de parasitologie-mycologie, présentées dans le Catalogue des analyses rares de la collégiale Anofel, et leur intérêt pour le diagnostic de ces infections.

Published
2020
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19. Selecting a multiplex PCR panel for accurate molecular diagnosis of intestinal protists: a comparative study of Allplex® (Seegene®), G-DiaParaTrio (Diagenode®), and RIDA®GENE (R-Biopharm®) assays and microscopic examination

Author

Nicolas Argy, Céline Nourrisson, Ahmed Aboubacar, Philippe Poirier, Stéphane Valot, Adrien Laude, Guillaume Desoubeaux, Christelle Pomares, Marie Machouart, Yohann Le Govic, Frédéric Dalle, Françoise Botterel, Nathalie Bourgeois, Estelle Cateau, Marion Leterrier, Patrice Le Pape, Florent Morio, and Sandrine Houze

Subjects
Infectious Diseases, Veterinary (miscellaneous), Insect Science, parasitic diseases, Animal Science and Zoology, Parasitology
Abstract

Commercial multiplex PCR assay panels were developed to overcome the limitations of microscopic examination for parasitological diagnosis on stool samples. However, given the increased supply of this diagnostic approach, these assays must be evaluated to position them in a diagnostic algorithm. Analytical performances of the multiplex PCR assay G-DiaParaTrio, Allplex® GI parasite and RIDA®GENE parasitic stool panel for detecting Blastocystis sp., Entamoeba histolytica, Giardia duodenalis, Cryptosporidium spp., Dientamoeba fragilis, and Cyclospora cayetanensis, were assessed through a retrospective comparative study on 184 stool samples initially sent for parasitological investigation. The composite reference method for parasitological diagnosis was microscopic observation and Entamoeba histolytica-specific adhesion detection when necessary. Multiplex PCR assays were performed on extracted DNA from each stool, following the manufacturer’s recommendations. Discrepant results with the composite reference method were investigated with species-specific PCR to approach a final parasitological diagnosis. Overall sensitivity/specificity for the multiplex PCR assays was 93.2%/100% for G-DiaParaTrio, 96.5%/98.3% for Allplex® GI parasite and 89.6%/98.3% for RIDA®GENE, whereas the composite reference method presented an overall sensitivity/specificity of 59.6%/99.8%. These results confirmed the added diagnostic value of the multiplex PCR approach for gastrointestinal protists. Nevertheless, the PCR procedure and the analytical performance for each protist of interest, variable depending on the multiplex PCR assay, must be considered when implementing a PCR-based diagnostic approach.

Published
2022
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20. Evaluation of a new multiplex PCR assay (ParaGENIE G-Amoeba Real-Time PCR kit) targeting Giardia intestinalis, Entamoeba histolytica and Entamoeba dispar/Entamoeba moshkovskii from stool specimens: evidence for the limited performances of microscopy-based approach for amoeba species identification

Author

Estelle Cateau, Nathalie Bourgeois, Marie Machouart, Florent Morio, Fakhri Jeddi, M. Gaboyard, Françoise Botterel, Stéphane Valot, Céline Nourrisson, Frédéric Dalle, Christelle Pomares, P. Le Pape, Adrien Laude, Guillaume Desoubeaux, Y. Le Govic, M. Leterrier, Nicolas Argy, Cibles et Médicaments des Infections et de l'Immunité (IICiMed), Université de Nantes - UFR des Sciences Pharmaceutiques et Biologiques, Université de Nantes (UN)-Université de Nantes (UN), Laboratoire de parasitologie mycologie (CHU de Dijon), Centre Hospitalier Universitaire de Dijon - Hôpital François Mitterrand (CHU Dijon), Laboratoire d'économie d'Orleans [2008-2011] (LEO), Université d'Orléans (UO)-Centre National de la Recherche Scientifique (CNRS), Université de Tours (UT), Services de Maladies Infectieuses et Tropicales [CHU Bichat], AP-HP - Hôpital Bichat - Claude Bernard [Paris], Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP), Laboratoire Microorganismes : Génome et Environnement (LMGE), Université Blaise Pascal - Clermont-Ferrand 2 (UBP)-Université d'Auvergne - Clermont-Ferrand I (UdA)-Centre National de la Recherche Scientifique (CNRS), Service de Parasitologie-Mycologie [CHRU Nancy], Centre Hospitalier Régional Universitaire de Nancy (CHRU Nancy), Groupe d'Étude des Interactions Hôte-Pathogène (GEIHP), Université d'Angers (UA), Service des Recherches Métallurgiques Appliquées (SRMA), Département des Matériaux pour le Nucléaire (DMN), CEA-Direction des Energies (ex-Direction de l'Energie Nucléaire) (CEA-DES (ex-DEN)), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Université Paris-Saclay-CEA-Direction des Energies (ex-Direction de l'Energie Nucléaire) (CEA-DES (ex-DEN)), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Université Paris-Saclay, Hôpital Henri Mondor, Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Hôpital Henri Mondor-Université Paris-Est Créteil Val-de-Marne - Paris 12 (UPEC UP12), Service de pharmacologie clinique et vigilances, Centre régional, de pharmacovigilance et de renseignement sur les médicaments, Hopital de la Milétrie, Poitiers, Microbiologie de l'Eau (MDE), Ecologie et biologie des interactions (EBI), Université de Poitiers-Centre National de la Recherche Scientifique (CNRS)-Université de Poitiers-Centre National de la Recherche Scientifique (CNRS), Laboratoire de Parasitologie-Mycologie (CHU de Poitiers), Centre hospitalier universitaire de Poitiers (CHU Poitiers), Service de Bactériologie-Hygiène, Institut Pasteur de Tunis, Réseau International des Instituts Pasteur (RIIP), Institut des Sciences Moléculaires (ISM), Université Montesquieu - Bordeaux 4-Université Sciences et Technologies - Bordeaux 1 (UB)-École Nationale Supérieure de Chimie et de Physique de Bordeaux (ENSCPB)-Institut de Chimie du CNRS (INC)-Centre National de la Recherche Scientifique (CNRS), Laboratoire de Parasitologie-Mycologie (Institut de biologie, CHU de Nantes), Centre hospitalier universitaire de Nantes (CHU Nantes), IICiMed - EA 1155, Université de Nantes (UN)-UFR Sciences et Techniques [Université de Nantes], Université de Nantes (UN)-UFR des Sciences Pharmaceutiques [Université de Nantes], Université de Nantes (UN), Laboratoire d'économie d'Orleans (LEO), Université de Tours, L’UNAM Université, Groupe d’Etude des Interactions Hôte-Pathogène, UPRES EA 3142, CEA-Direction de l'Energie Nucléaire (CEA-DEN), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Université Paris-Saclay-CEA-Direction de l'Energie Nucléaire (CEA-DEN), Assistance publique - Hôpitaux de Paris (AP-HP) (APHP)-Hôpital Henri Mondor-Université Paris-Est Créteil Val-de-Marne - Paris 12 (UPEC UP12), Statistique, Analyse et Modélisation Multidisciplinaire (SAmos-Marin Mersenne) (SAMM), Université Panthéon-Sorbonne (UP1), Université Montesquieu - Bordeaux 4-Université Sciences et Technologies - Bordeaux 1-École Nationale Supérieure de Chimie et de Physique de Bordeaux (ENSCPB)-Centre National de la Recherche Scientifique (CNRS), Géosciences Paris Sud (GEOPS), Université Paris-Sud - Paris 11 (UP11)-Centre National de la Recherche Scientifique (CNRS), Cibles et médicaments de l'infection, de l'immunité et du cancer (IICiMed), Université Montesquieu - Bordeaux 4-Université Sciences et Technologies - Bordeaux 1-École Nationale Supérieure de Chimie et de Physique de Bordeaux (ENSCPB)-Institut de Chimie du CNRS (INC)-Centre National de la Recherche Scientifique (CNRS), CHU Dijon, Biologie–Pharmacie, Pharmacie Centrale [CHU Nantes] (Pôle hospitalo-universitaire PHU7 - BPPC), Centre méditerranéen de médecine moléculaire (C3M), Université Nice Sophia Antipolis (... - 2019) (UNS), Université Côte d'Azur (UCA)-Université Côte d'Azur (UCA)-Institut National de la Santé et de la Recherche Médicale (INSERM), Laboratoire d'analyse et modélisation de systèmes pour l'aide à la décision (LAMSADE), Université Paris Dauphine-PSL-Centre National de la Recherche Scientifique (CNRS), Université de Poitiers-Centre National de la Recherche Scientifique (CNRS), Centre Hospitalier Départemental - Hôpital de La Roche-sur-Yon, CHU Montpellier, Centre Hospitalier Régional Universitaire [Montpellier] (CHRU Montpellier), Ademtech S.A., and Université Blaise Pascal - Clermont-Ferrand 2 (UBP)-Centre National de la Recherche Scientifique (CNRS)-Université d'Auvergne - Clermont-Ferrand I (UdA)

Subjects
Giardiasis, 0301 basic medicine, diagnosis, Entamoeba, Feces, Entamoeba dispar/Entamoeba moshkovskii, Giardia intestinalis, 0302 clinical medicine, fluids and secretions, Entamoeba moshkovskii, samples, [SDV.EE]Life Sciences [q-bio]/Ecology, environment, Amoeba species, Microscopy, parasitic infections, Entamoebiasis, biology, Entamoeba histolytica, General Medicine, 3. Good health, [SDV.MP]Life Sciences [q-bio]/Microbiology and Parasitology, Infectious Diseases, Real-time polymerase chain reaction, Stool, [SDV.MP.VIR]Life Sciences [q-bio]/Microbiology and Parasitology/Virology, antigen-detection, Microbiology (medical), food.ingredient, 030231 tropical medicine, 030106 microbiology, Real-Time Polymerase Chain Reaction, [SDV.MP.PRO]Life Sciences [q-bio]/Microbiology and Parasitology/Protistology, Microbiology, Amoeba (genus), 03 medical and health sciences, food, Species Specificity, Multiplex polymerase chain reaction, parasitic diseases, Humans, spp, dispar, Multiplex PCR, biology.organism_classification, [SDV.MP.BAC]Life Sciences [q-bio]/Microbiology and Parasitology/Bacteriology, Bacterial adhesin, amebiasis, Giardia lamblia, Multiplex Polymerase Chain Reaction, [SDV.MHEP]Life Sciences [q-bio]/Human health and pathology
Abstract

Objectives Besides the potential to identify a wide variety of gastrointestinal parasites, microscopy remains the reference standard in clinical microbiology for amoeba species identification and, especially when coupled with adhesin detection, to discriminate the pathogenic Entamoeba histolytica from its sister but non-pathogenic species Entamoeba dispar / Entamoeba moshkovskii . However, this approach is time-consuming, requires a high-level of expertise that can be jeopardized considering the low prevalence of gastrointestinal parasites in non-endemic countries. Here, we evaluated the CE-IVD-marked multiplex PCR (ParaGENIE G-Amoeba, Ademtech) targeting E. histolytica and E. dispar / E. moshkovskii and Giardia intestinalis . Methods This evaluation was performed blindly on a reference panel of 172 clinical stool samples collected prospectively from 12 laboratories and analysed using a standardized protocol relying on microscopy (and adhesin detection by ELISA for the detection of E. histolytica ) including G. intestinalis ( n = 37), various amoeba species ( n = 55) including E. dispar ( n = 15), E. histolytica ( n = 5), as well as 17 other gastrointestinal parasites ( n = 80), and negative samples ( n = 37). Results This new multiplex PCR assay offers fast and reliable results with appropriate sensitivity and specificity for the detection of G. intestinalis and E. dispar / E. moshkovskii from stools (89.7%/96.9% and 95%/100%, respectively). Detection rate and specificity were greatly improved by the PCR assay, highlighting several samples misidentified by microscopy, including false-negative and false-positive results for both E. dispar/E. moshkovskii and E. histolytica . Conclusion Given the clinical relevance of amoeba species identification, microbiologists should be aware of the limitations of using an algorithm relying on microscopy coupled with adhesin detection by ELISA.

Published
2018
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21. Is It Time to Include CT 'Reverse Halo Sign' and qPCR Targeting Mucorales in Serum to EORTC-MSG Criteria for the Diagnosis of Pulmonary Mucormycosis in Leukemia Patients ?

Author

Laurence Million, Denis Caillot, Marie Lorraine Chretien, Frédéric Dalle, Ingrid Lafon, Alexandre Payssot, Louise Basmaciyan, Stéphane Valot, Marc Sautour, Caroline Legouge, Service d'Hématologie Clinique (CHU de Dijon), Centre Hospitalier Universitaire de Dijon - Hôpital François Mitterrand (CHU Dijon), Lipides - Nutrition - Cancer (U866) (LNC), Université de Bourgogne (UB)-Institut National de la Santé et de la Recherche Médicale (INSERM)-AgroSup Dijon - Institut National Supérieur des Sciences Agronomiques, de l'Alimentation et de l'Environnement-Ecole Nationale Supérieure de Biologie Appliquée à la Nutrition et à l'Alimentation de Dijon (ENSBANA), Laboratoire d'Excellence : Lipoprotéines et Santé : prévention et Traitement des maladies Inflammatoires et du Cancer (LabEx LipSTIC), Institut National de la Recherche Agronomique (INRA)-Université Montpellier 2 - Sciences et Techniques (UM2)-Université Paris-Sud - Paris 11 (UP11)-École pratique des hautes études (EPHE)-Institut Gustave Roussy (IGR)-Centre Hospitalier Régional Universitaire de Nancy (CHRU Nancy)-Université de Bourgogne (UB)-Centre Hospitalier Universitaire de Dijon - Hôpital François Mitterrand (CHU Dijon)-Centre Régional de Lutte contre le cancer - Centre Georges-François Leclerc (CRLCC - CGFL)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Fédération Francophone de la Cancérologie Digestive, FFCD-AgroSup Dijon - Institut National Supérieur des Sciences Agronomiques, de l'Alimentation et de l'Environnement-Etablissement français du sang [Bourgogne-France-Comté] (EFS [Bourgogne-France-Comté])-Centre National de la Recherche Scientifique (CNRS)-Centre Hospitalier Régional Universitaire [Besançon] (CHRU Besançon)-Université de Franche-Comté (UFC)-Université de Montpellier (UM), Laboratoire de parasitologie mycologie (CHU de Dijon), Vin Aliment Microbiologie et Stress (VAlMiS), Procédés Alimentaires et Microbiologiques (PAM), Université de Bourgogne (UB)-AgroSup Dijon - Institut National Supérieur des Sciences Agronomiques, de l'Alimentation et de l'Environnement-Université de Bourgogne (UB)-AgroSup Dijon - Institut National Supérieur des Sciences Agronomiques, de l'Alimentation et de l'Environnement-Procédés Alimentaires et Microbiologiques [Dijon] (PAM), Université de Bourgogne (UB)-AgroSup Dijon - Institut National Supérieur des Sciences Agronomiques, de l'Alimentation et de l'Environnement-Université Bourgogne Franche-Comté [COMUE] (UBFC)-AgroSup Dijon - Institut National Supérieur des Sciences Agronomiques, de l'Alimentation et de l'Environnement-Université Bourgogne Franche-Comté [COMUE] (UBFC), Laboratoire Chrono-environnement - UFC (UMR 6249) (LCE), Université Bourgogne Franche-Comté [COMUE] (UBFC)-Centre National de la Recherche Scientifique (CNRS)-Université de Franche-Comté (UFC), Service de parasitologie et mycologie [CHU de Besançon], Centre Hospitalier Régional Universitaire [Besançon] (CHRU Besançon), Institut National de la Recherche Agronomique (INRA)-Université Montpellier 2 - Sciences et Techniques (UM2)-Université Paris-Sud - Paris 11 (UP11)-École pratique des hautes études (EPHE), Université Paris sciences et lettres (PSL)-Université Paris sciences et lettres (PSL)-Institut Gustave Roussy (IGR)-Centre Hospitalier Régional Universitaire de Nancy (CHRU Nancy)-Centre Hospitalier Régional Universitaire de Besançon (CHRU Besançon)-Université de Bourgogne (UB)-Centre Hospitalier Universitaire de Dijon - Hôpital François Mitterrand (CHU Dijon)-Centre Régional de Lutte contre le cancer Georges-François Leclerc [Dijon] (UNICANCER/CRLCC-CGFL), UNICANCER-UNICANCER-Institut National de la Santé et de la Recherche Médicale (INSERM)-Fédération Francophone de la Cancérologie Digestive, FFCD-Université de Montpellier (UM)-AgroSup Dijon - Institut National Supérieur des Sciences Agronomiques, de l'Alimentation et de l'Environnement-Etablissement français du sang [Bourgogne-Franche-Comté] (EFS [Bourgogne-Franche-Comté])-Centre National de la Recherche Scientifique (CNRS)-Université de Franche-Comté (UFC), Université Bourgogne Franche-Comté [COMUE] (UBFC)-Université Bourgogne Franche-Comté [COMUE] (UBFC), Laboratoire Chrono-environnement - CNRS - UBFC (UMR 6249) (LCE), Centre National de la Recherche Scientifique (CNRS)-Université de Franche-Comté (UFC), Service de parasitologie et mycologie [CHRU de Besançon], Centre Hospitalier Régional Universitaire de Besançon (CHRU Besançon), Centre Hospitalier Universitaire de Dijon - Hôpital François Mitterrand ( CHU Dijon ), Lipides - Nutrition - Cancer (U866) ( LNC ), Université de Bourgogne ( UB ) -Institut National de la Santé et de la Recherche Médicale ( INSERM ) -AgroSup Dijon - Institut National Supérieur des Sciences Agronomiques, de l'Alimentation et de l'Environnement-Ecole Nationale Supérieure de Biologie Appliquée à la Nutrition et à l'Alimentation de Dijon ( ENSBANA ), Laboratoire d'Excellence : Lipoprotéines et Santé : prévention et Traitement des maladies Inflammatoires et du Cancer ( LabEx LipSTIC ), Institut National de la Recherche Agronomique ( INRA ) -Université Montpellier 2 - Sciences et Techniques ( UM2 ) -Université Paris-Sud - Paris 11 ( UP11 ) -École pratique des hautes études ( EPHE ) -Institut Gustave Roussy ( IGR ) -Centre Hospitalier Régional Universitaire de Nancy ( CHRU Nancy ) -Université de Bourgogne ( UB ) -Centre Hospitalier Universitaire de Dijon - Hôpital François Mitterrand ( CHU Dijon ) -Centre Régional de Lutte contre le cancer - Centre Georges-François Leclerc ( CRLCC - CGFL ) -Institut National de la Santé et de la Recherche Médicale ( INSERM ) -Fédération Francophone de la Cancérologie Digestive, FFCD-AgroSup Dijon - Institut National Supérieur des Sciences Agronomiques, de l'Alimentation et de l'Environnement-Etablissement français du sang [Bourgogne-France-Comté] ( EFS [Bourgogne-France-Comté] ) -Centre National de la Recherche Scientifique ( CNRS ) -Centre Hospitalier Régional Universitaire [Besançon] ( CHRU Besançon ) -Université de Franche-Comté ( UFC ), Vin Aliment Microbiologie et Stress ( VAlMiS ), Procédés Alimentaires et Microbiologiques ( PAM ), Université de Bourgogne ( UB ) -AgroSup Dijon - Institut National Supérieur des Sciences Agronomiques, de l'Alimentation et de l'Environnement-Université de Bourgogne ( UB ) -AgroSup Dijon - Institut National Supérieur des Sciences Agronomiques, de l'Alimentation et de l'Environnement-Procédés Alimentaires et Microbiologiques [Dijon] ( PAM ), Université de Bourgogne ( UB ) -AgroSup Dijon - Institut National Supérieur des Sciences Agronomiques, de l'Alimentation et de l'Environnement-Université Bourgogne Franche-Comté ( UBFC ) -AgroSup Dijon - Institut National Supérieur des Sciences Agronomiques, de l'Alimentation et de l'Environnement-Université Bourgogne Franche-Comté ( UBFC ), Laboratoire Chrono-environnement ( LCE ), Université Bourgogne Franche-Comté ( UBFC ) -Centre National de la Recherche Scientifique ( CNRS ) -Université de Franche-Comté ( UFC ), Centre Hospitalier Régional Universitaire [Besançon] ( CHRU Besançon ), Institut National de la Santé et de la Recherche Médicale (INSERM)-Université de Bourgogne (UB)-Ecole Nationale Supérieure de Biologie Appliquée à la Nutrition et à l'Alimentation de Dijon (ENSBANA)-AgroSup Dijon - Institut National Supérieur des Sciences Agronomiques, de l'Alimentation et de l'Environnement, Université Paris sciences et lettres (PSL)-Université Paris sciences et lettres (PSL)-Institut Gustave Roussy (IGR)-Centre Hospitalier Régional Universitaire de Nancy (CHRU Nancy)-Université de Bourgogne (UB)-Centre Hospitalier Universitaire de Dijon - Hôpital François Mitterrand (CHU Dijon)-Centre Régional de Lutte contre le cancer Georges-François Leclerc [Dijon] (UNICANCER/CRLCC-CGFL), UNICANCER-UNICANCER-Institut National de la Santé et de la Recherche Médicale (INSERM)-Fédération Francophone de la Cancérologie Digestive, FFCD-AgroSup Dijon - Institut National Supérieur des Sciences Agronomiques, de l'Alimentation et de l'Environnement-Etablissement français du sang [Bourgogne-Franche-Comté] (EFS [Bourgogne-Franche-Comté])-Centre National de la Recherche Scientifique (CNRS)-Centre Hospitalier Régional Universitaire de Besançon (CHRU Besançon)-Université de Franche-Comté (UFC), Université Bourgogne Franche-Comté [COMUE] (UBFC)-Université Bourgogne Franche-Comté [COMUE] (UBFC)-Université de Montpellier (UM), Université Bourgogne Franche-Comté [COMUE] (UBFC)-Université de Bourgogne (UB)-AgroSup Dijon - Institut National Supérieur des Sciences Agronomiques, de l'Alimentation et de l'Environnement-Université Bourgogne Franche-Comté [COMUE] (UBFC)-AgroSup Dijon - Institut National Supérieur des Sciences Agronomiques, de l'Alimentation et de l'Environnement, Université de Franche-Comté (UFC), and Université Bourgogne Franche-Comté [COMUE] (UBFC)-Université Bourgogne Franche-Comté [COMUE] (UBFC)-Centre National de la Recherche Scientifique (CNRS)

Subjects
0301 basic medicine, Mucorales, Pathology, medicine.medical_specialty, 030106 microbiology, Computed tomography, law.invention, Reversed halo sign, 03 medical and health sciences, 0302 clinical medicine, law, Quantitative polymerase chain reaction, medicine, 030212 general & internal medicine, Pulmonary mucormycosis, Halo sign, Polymerase chain reaction, [SDV.MP.MYC]Life Sciences [q-bio]/Microbiology and Parasitology/Mycology, Leukemia, biology, medicine.diagnostic_test, business.industry, Brief Report, [ SDV.IDA ] Life Sciences [q-bio]/Food engineering, biology.organism_classification, medicine.disease, Early diagnosis, 3. Good health, Infectious Diseases, Real-time polymerase chain reaction, Oncology, medicine.symptom, Erratum, business
Abstract

In 23 leukemia patients with proven (n = 17) or possible (n = 6) pulmonary mucormycosis (PM), the presence of reversed halo sign on computed tomography was strongly associated with the positivity of quantitative polymerase chain reaction assays targeting Mucorales in the serum, confirming the value of these two tools for the diagnosis of PM in this setting.

Published
2016
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22. A misleading false-negative result of Pneumocystis real-time PCR assay due to a rare punctual mutation: A French multicenter study

Author

Laurence Pougnet, Sorya Belaz, Yann Pépino, Solène Le Gal, Lilia Hasseine, Christophe Hennequin, Stéphane Valot, Michèle Virmaux, Florence Robert-Gangneux, Jean-Philippe Bouchara, Mélanie Pitous, Eloise Lissillour, Thibaud Guillaud-Saumur, Gilles Nevez, Céline Damiani, Anne Totet, Dominique Toubas, Florent Morio, Paul Gueguen, Laboratoire de Parasitologie et Mycologiede [CHRU Brest], Centre Hospitalier Régional Universitaire de Brest (CHRU Brest), Groupe d'Étude des Interactions Hôte-Pathogène (GEIHP), Université d'Angers (UA), Service de Parasitologie-Mycologie [Rennes], Université de Rennes (UR)-Hôpital Pontchaillou-CHU Pontchaillou [Rennes], Institut de recherche en santé, environnement et travail (Irset), Université d'Angers (UA)-Université de Rennes (UR)-École des Hautes Études en Santé Publique [EHESP] (EHESP)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Structure Fédérative de Recherche en Biologie et Santé de Rennes ( Biosit : Biologie - Santé - Innovation Technologique ), Université de Brest (UBO), Périnatalité et Risques Toxiques - UMR INERIS_I 1 (PERITOX), Institut National de l'Environnement Industriel et des Risques (INERIS)-Université de Picardie Jules Verne (UPJV)-CHU Amiens-Picardie, Laboratoire de parasitologie et de mycologie médicales [CHU Amiens], CHU Amiens-Picardie, Génétique, génomique fonctionnelle et biotechnologies (UMR 1078) (GGB), EFS-Université de Brest (UBO)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Institut Brestois Santé Agro Matière (IBSAM), Laboratoire de Génétique Moléculaire et d'Histocompatibilité [Brest], Hôpital Morvan [Brest]-Centre Hospitalier Régional Universitaire de Brest (CHRU Brest), Matrice extracellulaire et dynamique cellulaire - UMR 7369 (MEDyC), Université de Reims Champagne-Ardenne (URCA)-SFR CAP Santé (Champagne-Ardenne Picardie Santé), Université de Reims Champagne-Ardenne (URCA)-Université de Reims Champagne-Ardenne (URCA)-Centre National de la Recherche Scientifique (CNRS), Centre Hospitalier Universitaire de Reims (CHU Reims), CHU Dijon, Centre Hospitalier Universitaire de Dijon - Hôpital François Mitterrand (CHU Dijon), Centre d'Immunologie et des Maladies Infectieuses (CIMI), Institut National de la Santé et de la Recherche Médicale (INSERM)-Sorbonne Université (SU)-Centre National de la Recherche Scientifique (CNRS), CHU Saint-Antoine [AP-HP], Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Sorbonne Université (SU), Centre hospitalier universitaire de Nantes (CHU Nantes), Centre Hospitalier Universitaire de Nice (CHU Nice), Agents infectieux, résistance et chimiothérapie - UR UPJV 4294 (AGIR ), Université de Picardie Jules Verne (UPJV)-CHU Amiens-Picardie, ERANet-LAC CAPRI-PC HID-0254, European Union, Université de Rennes 1 (UR1), Université de Rennes (UNIV-RENNES)-Université de Rennes (UNIV-RENNES)-Hôpital Pontchaillou-CHU Pontchaillou [Rennes], Structure Fédérative de Recherche en Biologie et Santé de Rennes ( Biosit : Biologie - Santé - Innovation Technologique )-Institut National de la Santé et de la Recherche Médicale (INSERM)-École des Hautes Études en Santé Publique [EHESP] (EHESP)-Université de Rennes 1 (UR1), Université de Rennes (UNIV-RENNES)-Université de Rennes (UNIV-RENNES)-Université d'Angers (UA), Université de Picardie Jules Verne (UPJV)-CHU Amiens-Picardie-Institut National de l'Environnement Industriel et des Risques, Institut Brestois Santé Agro Matière (IBSAM), Université de Brest (UBO)-Université de Brest (UBO)-EFS-Institut National de la Santé et de la Recherche Médicale (INSERM), Université de Reims Champagne-Ardenne (URCA)-Université de Picardie Jules Verne (UPJV)-Université de Reims Champagne-Ardenne (URCA)-Université de Picardie Jules Verne (UPJV)-Centre National de la Recherche Scientifique (CNRS), Service de Parasitologie - Mycologie [CHU Saint-Antoine], Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-CHU Saint-Antoine [AP-HP], Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Sorbonne Université (SU)-Sorbonne Université (SU), Centre d'Immunologie et de Maladies Infectieuses (CIMI), Centre National de la Recherche Scientifique (CNRS)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Université Pierre et Marie Curie - Paris 6 (UPMC), Departments of Medical Parasitology and Mycology, Université d'Angers (UA)-Université de Rennes 1 (UR1), Université de Rennes (UNIV-RENNES)-Université de Rennes (UNIV-RENNES)-École des Hautes Études en Santé Publique [EHESP] (EHESP)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Structure Fédérative de Recherche en Biologie et Santé de Rennes ( Biosit : Biologie - Santé - Innovation Technologique ), Centre Hospitalier Régional Universitaire de Brest (CHRU Brest)-Hôpital Morvan [Brest], SFR CAP Santé (Champagne-Ardenne Picardie Santé), Université de Reims Champagne-Ardenne (URCA)-Université de Picardie Jules Verne (UPJV)-Université de Reims Champagne-Ardenne (URCA)-Université de Picardie Jules Verne (UPJV)-Université de Reims Champagne-Ardenne (URCA)-Centre National de la Recherche Scientifique (CNRS), and Université Pierre et Marie Curie - Paris 6 (UPMC)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS)

Subjects
0301 basic medicine, Male, Pneumocystis pneumonia, Pneumocystis carinii, law.invention, Gene Frequency, law, Mutation frequency, Child, DNA, Fungal, False Negative Reactions, Polymerase chain reaction, Pneumocystis jirovecii, [SDV.EE]Life Sciences [q-bio]/Ecology, environment, Aged, 80 and over, Real time-PCR, biology, Pneumonia, Pneumocystis, General Medicine, Middle Aged, 3. Good health, PCP, Infectious Diseases, Real-time polymerase chain reaction, Molecular Diagnostic Techniques, Child, Preschool, Mutation (genetic algorithm), Female, France, Adult, Adolescent, 030106 microbiology, Real-Time Polymerase Chain Reaction, DNA, Mitochondrial, DNA, Ribosomal, 03 medical and health sciences, Young Adult, medicine, Humans, Point Mutation, Gene, TaqMan® MGB probe, Aged, Sequence Analysis, DNA, Ribosomal RNA, medicine.disease, biology.organism_classification, Virology, RNA, Ribosomal, mutation
Abstract

International audience; This article describes a previously unreported mutation at position 210 (C210T) of the mitochondrial large subunit ribosomal RNA (mtLSUrRNA) gene of Pneumocystis jirovecii, which led to a false-negative result of a real-time polymerase chain reaction (PCR) assay. Since the aforementioned real-time PCR assay is widely used in France, a French multicenter study was conducted to estimate the mutation frequency and its potential impact on the routine diagnosis of Pneumocystis pneumonia (PCP). Through analysis of data obtained from eight centers, the mutation frequency was estimated at 0.28%. This low frequency should not call into question the routine use of this PCR assay. Nonetheless, the occurrence of the false-negative PCR result provides arguments for maintaining microscopic techniques combined to PCR assays to achieve PCP diagnosis.

Published
2016
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23. Is real-time PCR-based diagnosis similar in performance to routine parasitological examination for the identification of Giardia intestinalis, Cryptosporidium parvum/Cryptosporidium hominis and Entamoeba histolytica from stool samples? Evaluation of a new commercial multiplex PCR assay and literature review

Author

Estelle Cateau, M. Leterrier, Frédéric Dalle, Adrien Laude, Nicolas Argy, P. Le Pape, Céline Nourrisson, Y. Le Govic, Guillaume Desoubeaux, Marie Machouart, Françoise Botterel, Nathalie Bourgeois, Florent Morio, Stéphane Valot, Christelle Pomares, Laboratoire d'économie d'Orleans ( LEO ), Université d'Orléans ( UO ) -Centre National de la Recherche Scientifique ( CNRS ), Université de Tours, Laboratoire Microorganismes : Génome et Environnement ( LMGE ), Université Blaise Pascal - Clermont-Ferrand 2 ( UBP ) -Centre National de la Recherche Scientifique ( CNRS ) -Université d'Auvergne - Clermont-Ferrand I ( UdA ), Relations Hôte-Environnement ( RHEM ), Université Henri Poincaré - Nancy 1 ( UHP ), Service de Parasitologie-Mycologie [CHRU Nancy], Centre Hospitalier Régional Universitaire de Nancy ( CHRU Nancy ), Agroécologie [Dijon], Institut National de la Recherche Agronomique ( INRA ) -Université de Bourgogne ( UB ) -AgroSup Dijon - Institut National Supérieur des Sciences Agronomiques, de l'Alimentation et de l'Environnement, Anofel Cryptosporidium National Network, Laboratoire de Parasitologie-Mycologie, Assistance publique - Hôpitaux de Paris (AP-HP)-Hôpital Henri Mondor-Université Paris-Est Créteil Val-de-Marne - Paris 12 ( UPEC UP12 ), Ecologie et biologie des interactions ( EBI ), Université de Poitiers-Centre National de la Recherche Scientifique ( CNRS ), Centre Hospitalier Regional Universitaire de Nantes, Géosciences Paris Sud ( GEOPS ), Université Paris-Sud - Paris 11 ( UP11 ) -Centre National de la Recherche Scientifique ( CNRS ), Laboratoire d'économie d'Orleans (LEO), Université d'Orléans (UO)-Centre National de la Recherche Scientifique (CNRS), Laboratoire de parasitologie mycologie (CHU de Dijon), Centre Hospitalier Universitaire de Dijon - Hôpital François Mitterrand (CHU Dijon), Services de Maladies Infectieuses et Tropicales [CHU Bichat], AP-HP - Hôpital Bichat - Claude Bernard [Paris], Laboratoire Microorganismes : Génome et Environnement (LMGE), Université Blaise Pascal - Clermont-Ferrand 2 (UBP)-Université d'Auvergne - Clermont-Ferrand I (UdA)-Centre National de la Recherche Scientifique (CNRS), Centre Hospitalier Régional Universitaire de Nancy (CHRU Nancy), Groupe d'Étude des Interactions Hôte-Pathogène (GEIHP), Université d'Angers (UA), Service des Recherches Métallurgiques Appliquées (SRMA), Département des Matériaux pour le Nucléaire (DMN), CEA-Direction des Energies (ex-Direction de l'Energie Nucléaire) (CEA-DES (ex-DEN)), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Université Paris-Saclay-CEA-Direction des Energies (ex-Direction de l'Energie Nucléaire) (CEA-DES (ex-DEN)), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Université Paris-Saclay, Hôpital Henri Mondor, Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Hôpital Henri Mondor-Université Paris-Est Créteil Val-de-Marne - Paris 12 (UPEC UP12), Centre hospitalier universitaire de Poitiers (CHU Poitiers), Service de Bactériologie-Hygiène, Géosciences Paris Sud (GEOPS), Université Paris-Sud - Paris 11 (UP11)-Centre National de la Recherche Scientifique (CNRS), IICiMed - EA 1155, Université de Nantes (UN)-UFR Sciences et Techniques [Université de Nantes], Université de Nantes (UN)-UFR des Sciences Pharmaceutiques [Université de Nantes], Université de Nantes (UN), Centre hospitalier universitaire de Nantes (CHU Nantes), CHU Dijon, Centre Hospitalier Régional Universitaire de Tours (CHRU Tours), Centre d’Etude des Pathologies Respiratoires (CEPR), UMR 1100 (CEPR), Université de Tours (UT)-Institut National de la Santé et de la Recherche Médicale (INSERM), Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP), Laboratoire de Parasitologie et Mycologie [CHU Clermont-Ferrand], CHU Gabriel Montpied [Clermont-Ferrand], CHU Clermont-Ferrand-CHU Clermont-Ferrand, Centre méditerranéen de médecine moléculaire (C3M), Université Nice Sophia Antipolis (1965 - 2019) (UNS), COMUE Université Côte d'Azur (2015-2019) (COMUE UCA)-COMUE Université Côte d'Azur (2015-2019) (COMUE UCA)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Université Côte d'Azur (UCA), Stress, Immunité, Pathogènes (SIMPA), Université de Lorraine (UL), Centre Hospitalier Universitaire d'Angers (CHU Angers), PRES Université Nantes Angers Le Mans (UNAM), Service de parasitologie [Mondor], Laboratoire de Parasitologie-Mycologie (CHU de Montpellier), Pôle Biologie-Pathologie [CHRU Montpellier], Centre Hospitalier Régional Universitaire [Montpellier] (CHRU Montpellier)-Centre Hospitalier Régional Universitaire [Montpellier] (CHRU Montpellier), Microbiologie de l'Eau (MDE), Ecologie et biologie des interactions (EBI), Université de Poitiers-Centre National de la Recherche Scientifique (CNRS)-Université de Poitiers-Centre National de la Recherche Scientifique (CNRS), Centre Hospitalier Départemental - Hôpital de La Roche-sur-Yon (CHD Vendée), Cibles et médicaments de l'infection, de l'immunité et du cancer (IICiMed), Université de Nantes - UFR des Sciences Pharmaceutiques et Biologiques, Université de Nantes (UN)-Université de Nantes (UN), Cibles et Médicaments des Infections et de l'Immunité (IICiMed), Université Blaise Pascal - Clermont-Ferrand 2 (UBP)-Centre National de la Recherche Scientifique (CNRS)-Université d'Auvergne - Clermont-Ferrand I (UdA), Laboratoire d'analyse et modélisation de systèmes pour l'aide à la décision (LAMSADE), Université Paris Dauphine-PSL, Université Paris sciences et lettres (PSL)-Université Paris sciences et lettres (PSL)-Centre National de la Recherche Scientifique (CNRS), Interactions et dynamique des environnements de surface (IDES), Université Paris-Sud - Paris 11 (UP11)-Institut national des sciences de l'Univers (INSU - CNRS)-Centre National de la Recherche Scientifique (CNRS), L’UNAM Université, Groupe d’Etude des Interactions Hôte-Pathogène, UPRES EA 3142, CEA-Direction de l'Energie Nucléaire (CEA-DEN), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Université Paris-Saclay-CEA-Direction de l'Energie Nucléaire (CEA-DEN), Assistance publique - Hôpitaux de Paris (AP-HP) (APHP)-Hôpital Henri Mondor-Université Paris-Est Créteil Val-de-Marne - Paris 12 (UPEC UP12), Statistique, Analyse et Modélisation Multidisciplinaire (SAmos-Marin Mersenne) (SAMM), Université Panthéon-Sorbonne (UP1), Université de Poitiers-Centre National de la Recherche Scientifique (CNRS), and Université de Tours (UT)

Subjects
0301 basic medicine, Microbiology (medical), 030231 tropical medicine, 030106 microbiology, Sensitivity and Specificity, [SDV.MP.PRO]Life Sciences [q-bio]/Microbiology and Parasitology/Protistology, Microbiology, 03 medical and health sciences, Entamoeba histolytica, Giardia intestinalis, Feces, 0302 clinical medicine, fluids and secretions, [ SDV.MHEP ] Life Sciences [q-bio]/Human health and pathology, Multiplex polymerase chain reaction, parasitic diseases, Humans, [SDV.MP.PAR]Life Sciences [q-bio]/Microbiology and Parasitology/Parasitology, Intestinal Diseases, Parasitic, stool, ComputingMilieux_MISCELLANEOUS, Cryptosporidium parvum, biology, [ SDV ] Life Sciences [q-bio], General Medicine, biology.organism_classification, DNA extraction, [SDV.MP.BAC]Life Sciences [q-bio]/Microbiology and Parasitology/Bacteriology, 3. Good health, Infectious Diseases, Real-time polymerase chain reaction, [SDV.MP]Life Sciences [q-bio]/Microbiology and Parasitology, Parasitology, Molecular Diagnostic Techniques, multiplex real-time PCR, microscopy, Giardia lamblia, Cryptosporidium hominis, Multiplex Polymerase Chain Reaction
Abstract

International audience; Microscopy is the reference standard for routine laboratory diagnosis in faecal parasitology but there is growing interest in alternative methods to overcome the limitations of microscopic examination, which is time-consuming and highly dependent on an operator's skills and expertise. Compared with microscopy, DNA detection by PCR is simple and can offer a better turnaround time. However, PCR performances remain difficult to assess as most studies have been conducted on a limited number of positive clinical samples and used in-house PCR methods. Our aim was to evaluate a new multiplex PCR assay (G-DiaParaTrio; Diagenode Diagnostics), targeting Giardia intestinalis, Cryptosporidium parvum/Cryptosporidium hominis and Entamoeba histolytica. To minimize the turnaround time, PCR was coupled with automated DNA extraction (QiaSymphony; Qiagen). The PCR assay was evaluated using a reference panel of 185 samples established by routine microscopic examination using a standardized protocol including Ziehl-Neelsen staining and adhesin detection by ELISA (E. histolytica II; TechLab). This panel, collected from 12 French parasitology laboratories, included 135 positive samples for G. intestinalis (n = 38), C. parvum/C. hominis (n = 26), E. histolytica (n = 5), 21 other gastrointestinal parasites, together with 50 negative samples. In all, the G-DiaParaTrio multiplex PCR assay identified 38 G. intestinalis, 25 C. parvum/C. hominis and five E. histolytica leading to sensitivity/specificity of 92%/100%, 96%/100% and 100%/100% for G. intestinalis, C. parvum/C. hominis and E. histolytica, respectively. This new multiplex PCR assay offers fast and reliable results, similar to microscopy-driven diagnosis for the detection of these gastrointestinal protozoa, allowing its implementation in routine clinical practice.

Published
2015
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