Performance evaluation of nine reference centers and comparison of DNA extraction protocols for effective surveillance of Leishmania-infected Phlebotomine sand flies: Basis for technical recommendations.
<h4>Background</h4>Leishmaniasis, caused by Leishmania protozoan parasites transmitted by Phlebotomine sand flies, is a significant public health concern in the Mediterranean basin. Effective monitoring of Leishmania-infected sand flies requires standardized tools for comparing their dis...
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Public Library of Science (PLoS)
2024-12-01
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| Series: | PLoS Neglected Tropical Diseases |
| Online Access: | https://doi.org/10.1371/journal.pntd.0012543 |
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| author | Jorian Prudhomme Aymeric Delabarre Bulent Alten Umut Berberoglu Eduardo Berriatua Gioia Bongiorno José Manuel Cristovao Maya Davidovich-Cohen Trentina Di Muccio Ozge Erisoz Kasap Eleonora Fiorentino Oscar D Kirstein Edwin Kniha Carla Maia Mesut Mungan Clara Muñoz-Hernández Muhammed Nalçaci Gizem Oguz Kaskan Yusuf Ozbel Seray Ozensoy Toz Ricardo Parreira Katharina Platzgummer Ceylan Polat José Risueño Liora Studentsky Gamze Varol Julia Walochnik Kardelen Yetişmiş Florence Robert-Gangneux |
| author_facet | Jorian Prudhomme Aymeric Delabarre Bulent Alten Umut Berberoglu Eduardo Berriatua Gioia Bongiorno José Manuel Cristovao Maya Davidovich-Cohen Trentina Di Muccio Ozge Erisoz Kasap Eleonora Fiorentino Oscar D Kirstein Edwin Kniha Carla Maia Mesut Mungan Clara Muñoz-Hernández Muhammed Nalçaci Gizem Oguz Kaskan Yusuf Ozbel Seray Ozensoy Toz Ricardo Parreira Katharina Platzgummer Ceylan Polat José Risueño Liora Studentsky Gamze Varol Julia Walochnik Kardelen Yetişmiş Florence Robert-Gangneux |
| author_sort | Jorian Prudhomme |
| collection | DOAJ |
| description | <h4>Background</h4>Leishmaniasis, caused by Leishmania protozoan parasites transmitted by Phlebotomine sand flies, is a significant public health concern in the Mediterranean basin. Effective monitoring of Leishmania-infected sand flies requires standardized tools for comparing their distribution and infection prevalence. Consistent quantitative real-time PCR (qPCR) parameters and efficient DNA extraction protocols are crucial for reliable results over time and across regions. However, the absence of standardized technical recommendations for Leishmania DNA detection hinders effective surveillance. This study aimed to compare different DNA extraction protocols and conduct a qPCR-based External Quality Assessment (EQA) through a multicenter study involving nine reference laboratories, with a focus on optimizing Leishmania DNA detection in sand fly.<h4>Methodology/principal findings</h4>EQA samples consisted of Leishmania infantum and L. major species, at concentrations ranging from 101 to 104 parasites/mL. All but one center detected all concentrations, demonstrating strong diagnostic proficiency. The ability to detect low concentrations highlighted the robustness of the qPCR assay used, though variations in Cq values indicated differences in sensitivity related to technical capabilities or DNA extraction kit performance. A comparative analysis of seven DNA extraction methods identified the EZ1 DSP Virus Kit and QIAamp DNA mini-kit as the most efficient, supporting their use in standardized protocols. The study also assessed the effects of lyophilization and shipment conditions, showing no significant compromise in Leishmania detection despite slight variations in Cq values. Experimentally infected sand flies were included to simulate field conditions, and all centers successfully detected positive samples with varying Cq values, probably reflecting differences in infection load.<h4>Conclusion and significance</h4>This study emphasizes the importance of standardized DNA extraction protocols and continuous quality assurance for accurate Leishmania DNA detection. The results highlight the superior performance of certain extraction kits and the need for ongoing technical training, essential for reliable leishmaniasis surveillance, particularly in field settings with low infection densities. |
| format | Article |
| id | doaj-art-10499ecff3e4450780c836b1da5d2ece |
| institution | Kabale University |
| issn | 1935-2727 1935-2735 |
| language | English |
| publishDate | 2024-12-01 |
| publisher | Public Library of Science (PLoS) |
| record_format | Article |
| series | PLoS Neglected Tropical Diseases |
| spelling | doaj-art-10499ecff3e4450780c836b1da5d2ece2025-01-29T05:31:04ZengPublic Library of Science (PLoS)PLoS Neglected Tropical Diseases1935-27271935-27352024-12-011812e001254310.1371/journal.pntd.0012543Performance evaluation of nine reference centers and comparison of DNA extraction protocols for effective surveillance of Leishmania-infected Phlebotomine sand flies: Basis for technical recommendations.Jorian PrudhommeAymeric DelabarreBulent AltenUmut BerberogluEduardo BerriatuaGioia BongiornoJosé Manuel CristovaoMaya Davidovich-CohenTrentina Di MuccioOzge Erisoz KasapEleonora FiorentinoOscar D KirsteinEdwin KnihaCarla MaiaMesut MunganClara Muñoz-HernándezMuhammed NalçaciGizem Oguz KaskanYusuf OzbelSeray Ozensoy TozRicardo ParreiraKatharina PlatzgummerCeylan PolatJosé RisueñoLiora StudentskyGamze VarolJulia WalochnikKardelen YetişmişFlorence Robert-Gangneux<h4>Background</h4>Leishmaniasis, caused by Leishmania protozoan parasites transmitted by Phlebotomine sand flies, is a significant public health concern in the Mediterranean basin. Effective monitoring of Leishmania-infected sand flies requires standardized tools for comparing their distribution and infection prevalence. Consistent quantitative real-time PCR (qPCR) parameters and efficient DNA extraction protocols are crucial for reliable results over time and across regions. However, the absence of standardized technical recommendations for Leishmania DNA detection hinders effective surveillance. This study aimed to compare different DNA extraction protocols and conduct a qPCR-based External Quality Assessment (EQA) through a multicenter study involving nine reference laboratories, with a focus on optimizing Leishmania DNA detection in sand fly.<h4>Methodology/principal findings</h4>EQA samples consisted of Leishmania infantum and L. major species, at concentrations ranging from 101 to 104 parasites/mL. All but one center detected all concentrations, demonstrating strong diagnostic proficiency. The ability to detect low concentrations highlighted the robustness of the qPCR assay used, though variations in Cq values indicated differences in sensitivity related to technical capabilities or DNA extraction kit performance. A comparative analysis of seven DNA extraction methods identified the EZ1 DSP Virus Kit and QIAamp DNA mini-kit as the most efficient, supporting their use in standardized protocols. The study also assessed the effects of lyophilization and shipment conditions, showing no significant compromise in Leishmania detection despite slight variations in Cq values. Experimentally infected sand flies were included to simulate field conditions, and all centers successfully detected positive samples with varying Cq values, probably reflecting differences in infection load.<h4>Conclusion and significance</h4>This study emphasizes the importance of standardized DNA extraction protocols and continuous quality assurance for accurate Leishmania DNA detection. The results highlight the superior performance of certain extraction kits and the need for ongoing technical training, essential for reliable leishmaniasis surveillance, particularly in field settings with low infection densities.https://doi.org/10.1371/journal.pntd.0012543 |
| spellingShingle | Jorian Prudhomme Aymeric Delabarre Bulent Alten Umut Berberoglu Eduardo Berriatua Gioia Bongiorno José Manuel Cristovao Maya Davidovich-Cohen Trentina Di Muccio Ozge Erisoz Kasap Eleonora Fiorentino Oscar D Kirstein Edwin Kniha Carla Maia Mesut Mungan Clara Muñoz-Hernández Muhammed Nalçaci Gizem Oguz Kaskan Yusuf Ozbel Seray Ozensoy Toz Ricardo Parreira Katharina Platzgummer Ceylan Polat José Risueño Liora Studentsky Gamze Varol Julia Walochnik Kardelen Yetişmiş Florence Robert-Gangneux Performance evaluation of nine reference centers and comparison of DNA extraction protocols for effective surveillance of Leishmania-infected Phlebotomine sand flies: Basis for technical recommendations. PLoS Neglected Tropical Diseases |
| title | Performance evaluation of nine reference centers and comparison of DNA extraction protocols for effective surveillance of Leishmania-infected Phlebotomine sand flies: Basis for technical recommendations. |
| title_full | Performance evaluation of nine reference centers and comparison of DNA extraction protocols for effective surveillance of Leishmania-infected Phlebotomine sand flies: Basis for technical recommendations. |
| title_fullStr | Performance evaluation of nine reference centers and comparison of DNA extraction protocols for effective surveillance of Leishmania-infected Phlebotomine sand flies: Basis for technical recommendations. |
| title_full_unstemmed | Performance evaluation of nine reference centers and comparison of DNA extraction protocols for effective surveillance of Leishmania-infected Phlebotomine sand flies: Basis for technical recommendations. |
| title_short | Performance evaluation of nine reference centers and comparison of DNA extraction protocols for effective surveillance of Leishmania-infected Phlebotomine sand flies: Basis for technical recommendations. |
| title_sort | performance evaluation of nine reference centers and comparison of dna extraction protocols for effective surveillance of leishmania infected phlebotomine sand flies basis for technical recommendations |
| url | https://doi.org/10.1371/journal.pntd.0012543 |
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