Development of a New Internally Controlled One-Step Real-Time RT-PCR for the Molecular Detection of Enterovirus A71 in Africa and Madagascar
Enterovirus A71 (EV-A71) is a leading cause of hand-foot-and-mouth disease (HFMD) and can be associated with severe neurological complications. EV-A71 strains can be classified into seven genogroups, A-H, on the basis of the VP1 capsid protein gene sequence. Genogroup A includes the prototype strain...
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| Format: | Article |
| Language: | English |
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Frontiers Media S.A.
2020-08-01
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| Series: | Frontiers in Microbiology |
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| Online Access: | https://www.frontiersin.org/article/10.3389/fmicb.2020.01907/full |
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| author | Romain Volle Romain Volle Marie-Line Joffret Marie-Line Joffret Marie-Line Joffret Kader Ndiaye Maria Dolores Fernandez-Garcia Richter Razafindratsimandresy Jean-Michel Heraud Dorra Rezig Serge Alain Sadeuh-Mba Leila Boulahbal-Anes Mohamed Seghier Jagadish M. Deshpandeh Maël Bessaud Maël Bessaud Maël Bessaud Francis Delpeyroux Francis Delpeyroux |
| author_facet | Romain Volle Romain Volle Marie-Line Joffret Marie-Line Joffret Marie-Line Joffret Kader Ndiaye Maria Dolores Fernandez-Garcia Richter Razafindratsimandresy Jean-Michel Heraud Dorra Rezig Serge Alain Sadeuh-Mba Leila Boulahbal-Anes Mohamed Seghier Jagadish M. Deshpandeh Maël Bessaud Maël Bessaud Maël Bessaud Francis Delpeyroux Francis Delpeyroux |
| author_sort | Romain Volle |
| collection | DOAJ |
| description | Enterovirus A71 (EV-A71) is a leading cause of hand-foot-and-mouth disease (HFMD) and can be associated with severe neurological complications. EV-A71 strains can be classified into seven genogroups, A-H, on the basis of the VP1 capsid protein gene sequence. Genogroup A includes the prototype strain; genogroups B and C are responsible of major outbreaks worldwide, but little is known about the others, particularly genogroups E and F, which have been recently identified in Africa and Madagascar, respectively. The circulation of EV-A71 in the African region is poorly known and probably underestimated. A rapid and specific assay for detecting all genogroups of EV-A71 is required. In this study, we developed a real-time RT-PCR assay with a competitive internal control (IC). The primers and TaqMan probe specifically target the genomic region encoding the VP1 capsid protein. Diverse EV-A71 RNAs were successfully amplified from the genogroups A, B, C, D, E, and F, with similar sensitivity and robust reproducibility. Neither cross reaction with other EVs nor major interference with the competitive IC was detected. Experimentally spiked stool and plasma specimens provided consistent and reproducible results, and validated the usefulness of the IC for demonstrating the presence of PCR inhibitors in samples. The analysis in an African laboratories network of 1889 untyped enterovirus isolates detected 15 EV-A71 of different genogroups. This specific real-time RT-PCR assay provides a robust and sensitive method for the detection of EV-A71 in biological specimens and for the epidemiological monitoring of EV-A71 including its recently discovered genogroups. |
| first_indexed | 2024-12-20T22:41:30Z |
| format | Article |
| id | doaj.art-63e6bc7f97b2487fbf7103045a0e02aa |
| institution | Directory Open Access Journal |
| issn | 1664-302X |
| language | English |
| last_indexed | 2024-12-20T22:41:30Z |
| publishDate | 2020-08-01 |
| publisher | Frontiers Media S.A. |
| record_format | Article |
| series | Frontiers in Microbiology |
| spelling | doaj.art-63e6bc7f97b2487fbf7103045a0e02aa2022-12-21T19:24:27ZengFrontiers Media S.A.Frontiers in Microbiology1664-302X2020-08-011110.3389/fmicb.2020.01907564076Development of a New Internally Controlled One-Step Real-Time RT-PCR for the Molecular Detection of Enterovirus A71 in Africa and MadagascarRomain Volle0Romain Volle1Marie-Line Joffret2Marie-Line Joffret3Marie-Line Joffret4Kader Ndiaye5Maria Dolores Fernandez-Garcia6Richter Razafindratsimandresy7Jean-Michel Heraud8Dorra Rezig9Serge Alain Sadeuh-Mba10Leila Boulahbal-Anes11Mohamed Seghier12Jagadish M. Deshpandeh13Maël Bessaud14Maël Bessaud15Maël Bessaud16Francis Delpeyroux17Francis Delpeyroux18Institut Pasteur, Unité de Biologie des Virus Entériques, Paris, FranceINSERM U994, Institut National de Santé et de La Recherche Médicale, Paris, FranceInstitut Pasteur, Unité de Biologie des Virus Entériques, Paris, FranceINSERM U994, Institut National de Santé et de La Recherche Médicale, Paris, FranceInstitut Pasteur, Viral Populations and Pathogenesis, Paris, FranceInstitut Pasteur de Dakar, Dakar, SenegalInstitut Pasteur de Dakar, Dakar, SenegalInstitut Pasteur de Madagascar, Antananarivo, MadagascarInstitut Pasteur de Madagascar, Antananarivo, MadagascarInstitut Pasteur de Tunis, Tunis, TunisiaCentre Pasteur du Cameroun, Yaoundé, CameroonInstitut Pasteur d’Algérie, Algiers, AlgeriaInstitut Pasteur d’Algérie, Algiers, AlgeriaNational Institute of Virology, Indian Council of Medical Research (ICMR), Mumbai, IndiaInstitut Pasteur, Unité de Biologie des Virus Entériques, Paris, FranceINSERM U994, Institut National de Santé et de La Recherche Médicale, Paris, FranceInstitut Pasteur, Viral Populations and Pathogenesis, Paris, FranceInstitut Pasteur, Unité de Biologie des Virus Entériques, Paris, FranceINSERM U994, Institut National de Santé et de La Recherche Médicale, Paris, FranceEnterovirus A71 (EV-A71) is a leading cause of hand-foot-and-mouth disease (HFMD) and can be associated with severe neurological complications. EV-A71 strains can be classified into seven genogroups, A-H, on the basis of the VP1 capsid protein gene sequence. Genogroup A includes the prototype strain; genogroups B and C are responsible of major outbreaks worldwide, but little is known about the others, particularly genogroups E and F, which have been recently identified in Africa and Madagascar, respectively. The circulation of EV-A71 in the African region is poorly known and probably underestimated. A rapid and specific assay for detecting all genogroups of EV-A71 is required. In this study, we developed a real-time RT-PCR assay with a competitive internal control (IC). The primers and TaqMan probe specifically target the genomic region encoding the VP1 capsid protein. Diverse EV-A71 RNAs were successfully amplified from the genogroups A, B, C, D, E, and F, with similar sensitivity and robust reproducibility. Neither cross reaction with other EVs nor major interference with the competitive IC was detected. Experimentally spiked stool and plasma specimens provided consistent and reproducible results, and validated the usefulness of the IC for demonstrating the presence of PCR inhibitors in samples. The analysis in an African laboratories network of 1889 untyped enterovirus isolates detected 15 EV-A71 of different genogroups. This specific real-time RT-PCR assay provides a robust and sensitive method for the detection of EV-A71 in biological specimens and for the epidemiological monitoring of EV-A71 including its recently discovered genogroups.https://www.frontiersin.org/article/10.3389/fmicb.2020.01907/fullenterovirus 71EV-A71HFMDnew genogroupsmolecular detection |
| spellingShingle | Romain Volle Romain Volle Marie-Line Joffret Marie-Line Joffret Marie-Line Joffret Kader Ndiaye Maria Dolores Fernandez-Garcia Richter Razafindratsimandresy Jean-Michel Heraud Dorra Rezig Serge Alain Sadeuh-Mba Leila Boulahbal-Anes Mohamed Seghier Jagadish M. Deshpandeh Maël Bessaud Maël Bessaud Maël Bessaud Francis Delpeyroux Francis Delpeyroux Development of a New Internally Controlled One-Step Real-Time RT-PCR for the Molecular Detection of Enterovirus A71 in Africa and Madagascar Frontiers in Microbiology enterovirus 71 EV-A71 HFMD new genogroups molecular detection |
| title | Development of a New Internally Controlled One-Step Real-Time RT-PCR for the Molecular Detection of Enterovirus A71 in Africa and Madagascar |
| title_full | Development of a New Internally Controlled One-Step Real-Time RT-PCR for the Molecular Detection of Enterovirus A71 in Africa and Madagascar |
| title_fullStr | Development of a New Internally Controlled One-Step Real-Time RT-PCR for the Molecular Detection of Enterovirus A71 in Africa and Madagascar |
| title_full_unstemmed | Development of a New Internally Controlled One-Step Real-Time RT-PCR for the Molecular Detection of Enterovirus A71 in Africa and Madagascar |
| title_short | Development of a New Internally Controlled One-Step Real-Time RT-PCR for the Molecular Detection of Enterovirus A71 in Africa and Madagascar |
| title_sort | development of a new internally controlled one step real time rt pcr for the molecular detection of enterovirus a71 in africa and madagascar |
| topic | enterovirus 71 EV-A71 HFMD new genogroups molecular detection |
| url | https://www.frontiersin.org/article/10.3389/fmicb.2020.01907/full |
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