Development of a ptp2-LAMP assay for the specific and sensitive detection of Nosema apis and its comparison with ptp3-LAMP for the detection of Nosema ceranae, in a region endemic for both microsporidium pathogens of the Western honey bee
The Western honey bee plays a pivotal role in global food security as the primary commercial pollinator. The microsporidian pathogens Nosema apis and Nosema ceranae infect the bee midgut, causing nosemosis, a debilitating infectious disease that results in considerable economic losses in apiculture....
| Autores principales: | , , , , |
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| Formato: | info:ar-repo/semantics/artículo |
| Lenguaje: | Inglés |
| Publicado: |
Elsevier
2025
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| Materias: | |
| Acceso en línea: | http://hdl.handle.net/20.500.12123/22342 https://www.sciencedirect.com/science/article/abs/pii/S0020751925000633 https://doi.org/10.1016/j.ijpara.2025.04.001 |
| _version_ | 1855038558091345920 |
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| author | Lannutti, Lucas Gisder, Sebastian Florin-Christensen, Monica Genersch, Elke Schnittger, Leonhard |
| author_browse | Florin-Christensen, Monica Genersch, Elke Gisder, Sebastian Lannutti, Lucas Schnittger, Leonhard |
| author_facet | Lannutti, Lucas Gisder, Sebastian Florin-Christensen, Monica Genersch, Elke Schnittger, Leonhard |
| author_sort | Lannutti, Lucas |
| collection | INTA Digital |
| description | The Western honey bee plays a pivotal role in global food security as the primary commercial pollinator. The microsporidian pathogens Nosema apis and Nosema ceranae infect the bee midgut, causing nosemosis, a debilitating infectious disease that results in considerable economic losses in apiculture. Traditionally, Nosema spp. infection is diagnosed by microscopic detection and quantification of spores. However, only molecular diagnostics allow differentiation between N. apis and N. ceranae. Loop-mediated isothermal amplification (LAMP) is a rapid, highly specific, and sensitive DNA detection method. The present study aimed to develop a LAMP protocol for N. apis based on the species-specific single copy polar tube protein 2 (ptp2) gene, and to analyze and compare its diagnostic performance with the previously developed polar tube protein 3 (ptp3) gene-based LAMP protocol for N. ceranae. The ptp2- and ptp3-LAMP assays specifically identified N. apis and N. ceranae, respectively. Their analytical sensitivity was tested using serial dilutions of plasmid and genomic DNA, demonstrating that ptp2- and ptp3-LAMP consistently detected down to 103 ptp2 and 104 ptp3-gene copies, respectively. Amplification was verified by agarose gel electrophoresis (conventional format), and by a change from pink to yellow color after addition of a suitable dye (colorimetric format). The ptp2- and ptp3-LAMP assays and a reference duplex PCR were applied to a panel of field samples (n = 55) from a region endemic for both Nosema spp. Conventional and colorimetric ptp2-LAMP showed an almost perfect test agreement (kappa value > 0.81) compared with duplex PCR. Conventional and colorimetric ptp3-LAMP assays showed a substantial (kappa value > 0.60) and almost perfect test agreement (kappa value > 0.81), respectively. The ptp2- and ptp3-LAMP assays provide excellent performance, ease of implementation, cost savings, and rapid execution, making them ideal choices for molecular detection and differentiation of N. apis and N. ceranae. |
| format | info:ar-repo/semantics/artículo |
| id | INTA22342 |
| institution | Instituto Nacional de Tecnología Agropecuaria (INTA -Argentina) |
| language | Inglés |
| publishDate | 2025 |
| publishDateRange | 2025 |
| publishDateSort | 2025 |
| publisher | Elsevier |
| publisherStr | Elsevier |
| record_format | dspace |
| spelling | INTA223422025-05-20T10:04:06Z Development of a ptp2-LAMP assay for the specific and sensitive detection of Nosema apis and its comparison with ptp3-LAMP for the detection of Nosema ceranae, in a region endemic for both microsporidium pathogens of the Western honey bee Lannutti, Lucas Gisder, Sebastian Florin-Christensen, Monica Genersch, Elke Schnittger, Leonhard Nosema apis Apis mellifera Honey Bees Microsporidiosis PCR Diagnostic Techniques Parasitology Abeja Melífera Técnica de Diagnóstico Parasitología Nosema ceranae The Western honey bee plays a pivotal role in global food security as the primary commercial pollinator. The microsporidian pathogens Nosema apis and Nosema ceranae infect the bee midgut, causing nosemosis, a debilitating infectious disease that results in considerable economic losses in apiculture. Traditionally, Nosema spp. infection is diagnosed by microscopic detection and quantification of spores. However, only molecular diagnostics allow differentiation between N. apis and N. ceranae. Loop-mediated isothermal amplification (LAMP) is a rapid, highly specific, and sensitive DNA detection method. The present study aimed to develop a LAMP protocol for N. apis based on the species-specific single copy polar tube protein 2 (ptp2) gene, and to analyze and compare its diagnostic performance with the previously developed polar tube protein 3 (ptp3) gene-based LAMP protocol for N. ceranae. The ptp2- and ptp3-LAMP assays specifically identified N. apis and N. ceranae, respectively. Their analytical sensitivity was tested using serial dilutions of plasmid and genomic DNA, demonstrating that ptp2- and ptp3-LAMP consistently detected down to 103 ptp2 and 104 ptp3-gene copies, respectively. Amplification was verified by agarose gel electrophoresis (conventional format), and by a change from pink to yellow color after addition of a suitable dye (colorimetric format). The ptp2- and ptp3-LAMP assays and a reference duplex PCR were applied to a panel of field samples (n = 55) from a region endemic for both Nosema spp. Conventional and colorimetric ptp2-LAMP showed an almost perfect test agreement (kappa value > 0.81) compared with duplex PCR. Conventional and colorimetric ptp3-LAMP assays showed a substantial (kappa value > 0.60) and almost perfect test agreement (kappa value > 0.81), respectively. The ptp2- and ptp3-LAMP assays provide excellent performance, ease of implementation, cost savings, and rapid execution, making them ideal choices for molecular detection and differentiation of N. apis and N. ceranae. Instituto de Patobiología Fil: Lannutti, Lucas. Universidad de Morón. Escuela Superior de Ciencias Exactas y Naturales; Argentina Fil: Lannutti, Lucas. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina Fil: Lannutti, Lucas. Institute for Bee Research. Department of Molecular Microbiology and Bee Diseases; Alemania Fil: Gisder, Sebastian. Institute for Bee Research. Department of Molecular Microbiology and Bee Diseases; Alemania Fil: Florin-Christensen, Monica. Universidad de Morón. Escuela Superior de Ciencias Exactas y Naturales; Argentina Fil: Florin-Christensen, Monica. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina Fil: Florin-Christensen, Monica. Instituto Nacional de Tecnología Agropecuaria (INTA). Instituto de Patobiología Veterinaria; Argentina Fil: Genersch, Elke. Institute for Bee Research. Department of Molecular Microbiology and Bee Diseases; Alemania Fil: Schnittger, Leonhard. Universidad de Morón. Escuela Superior de Ciencias Exactas y Naturales; Argentina Fil: Schnittger, Leonhard. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina Fil: Schnittger, Leonhard. Instituto Nacional de Tecnología Agropecuaria (INTA). Instituto de Patobiología Veterinaria; Argentina 2025-05-20T09:52:50Z 2025-05-20T09:52:50Z 2025 info:ar-repo/semantics/artículo info:eu-repo/semantics/article info:eu-repo/semantics/publishedVersion http://hdl.handle.net/20.500.12123/22342 https://www.sciencedirect.com/science/article/abs/pii/S0020751925000633 1879-0135 https://doi.org/10.1016/j.ijpara.2025.04.001 eng info:eu-repograntAgreement/INTA/2019-PE-E1-I017-001, Desarrollo del sector apícola organizado, sustentable y competitivo info:eu-repograntAgreement/INTA/2023-PE-L01-I069, Aportes al desarrollo sostenible de la apicultura argentina info:eu-repo/semantics/restrictedAccess http://creativecommons.org/licenses/by-nc-sa/4.0/ Creative Commons Attribution-NonCommercial-ShareAlike 4.0 International (CC BY-NC-SA 4.0) application/pdf Elsevier International Journal for Parasitology : 1-12 (Available online 5 April 2025) |
| spellingShingle | Nosema apis Apis mellifera Honey Bees Microsporidiosis PCR Diagnostic Techniques Parasitology Abeja Melífera Técnica de Diagnóstico Parasitología Nosema ceranae Lannutti, Lucas Gisder, Sebastian Florin-Christensen, Monica Genersch, Elke Schnittger, Leonhard Development of a ptp2-LAMP assay for the specific and sensitive detection of Nosema apis and its comparison with ptp3-LAMP for the detection of Nosema ceranae, in a region endemic for both microsporidium pathogens of the Western honey bee |
| title | Development of a ptp2-LAMP assay for the specific and sensitive detection of Nosema apis and its comparison with ptp3-LAMP for the detection of Nosema ceranae, in a region endemic for both microsporidium pathogens of the Western honey bee |
| title_full | Development of a ptp2-LAMP assay for the specific and sensitive detection of Nosema apis and its comparison with ptp3-LAMP for the detection of Nosema ceranae, in a region endemic for both microsporidium pathogens of the Western honey bee |
| title_fullStr | Development of a ptp2-LAMP assay for the specific and sensitive detection of Nosema apis and its comparison with ptp3-LAMP for the detection of Nosema ceranae, in a region endemic for both microsporidium pathogens of the Western honey bee |
| title_full_unstemmed | Development of a ptp2-LAMP assay for the specific and sensitive detection of Nosema apis and its comparison with ptp3-LAMP for the detection of Nosema ceranae, in a region endemic for both microsporidium pathogens of the Western honey bee |
| title_short | Development of a ptp2-LAMP assay for the specific and sensitive detection of Nosema apis and its comparison with ptp3-LAMP for the detection of Nosema ceranae, in a region endemic for both microsporidium pathogens of the Western honey bee |
| title_sort | development of a ptp2 lamp assay for the specific and sensitive detection of nosema apis and its comparison with ptp3 lamp for the detection of nosema ceranae in a region endemic for both microsporidium pathogens of the western honey bee |
| topic | Nosema apis Apis mellifera Honey Bees Microsporidiosis PCR Diagnostic Techniques Parasitology Abeja Melífera Técnica de Diagnóstico Parasitología Nosema ceranae |
| url | http://hdl.handle.net/20.500.12123/22342 https://www.sciencedirect.com/science/article/abs/pii/S0020751925000633 https://doi.org/10.1016/j.ijpara.2025.04.001 |
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