Rapid high throughput SYBR green assay for identifying the malaria vectors Anopheles arabiensis, Anopheles coluzzii and Anopheles gambiae s.s. Giles
| Autor: | Dimitra Pipini, Helen Pates Jamet, Louis Kouadio N'Dri, Samuel K. Dadzie, Arjen E Van’t Hof, Joseph Chabi, Takashi Suzuki, Dora Okyere, Melinda P. Hadi, Alex Kojo Datsomor, Dziedzom K. de Souza, Harun Njoroge |
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| Jazyk: | angličtina |
| Rok vydání: | 2018 |
| Předmět: |
0301 basic medicine
Species complex Science Anopheles gambiae 030231 tropical medicine Zoology Mosquito Vectors Diamines Ghana Polymerase Chain Reaction Sensitivity and Specificity law.invention 03 medical and health sciences 0302 clinical medicine Species Specificity Sensu law Anopheles parasitic diseases Animals Benzothiazoles Organic Chemicals Malaria vector Polymerase chain reaction Fluorescent Dyes 030304 developmental biology Hybrid Genetics 0303 health sciences Multidisciplinary biology Reproducibility of Results DNA Amplicon biology.organism_classification Kenya Malaria 3. Good health 030104 developmental biology Molecular Diagnostic Techniques Vector (epidemiology) Quinolines Medicine Anopheles coluzzii geographic locations |
| Zdroj: | PLoS ONE, Vol 14, Iss 4, p e0215669 (2019) |
| DOI: | 10.1101/448910 |
| Popis: | The Anopheles gambiae sensu lato species complex consists of a number of cryptic species with different habitats and behaviours. These morphologically indistinct species are identified by chromosome banding and molecular diagnostic techniques which are still under improvement even though the current SINE method for identification between An. coluzzii and An. gambiae works reliably. This study describes a refinement of the SINE method to increase sensitivity and high throughput method for the identification of both species and An. arabiensis using amplicon dissociation characteristics.Field collected samples, laboratory reared colonies and crossed specimens of the two species were used for the design of the protocol. An. gambiae, An. coluzzii, and hybrids of the two species were provided by the insectary of Vestergaard-NMIMR Vector Labs at the Noguchi Memorial Institute for Medical Research (Ghana) and An. arabiensis from Kenya. Samples were first characterised using conventional SINE PCR method, and further assayed using SYBR green, an intercalating fluorescent dye.The three species and hybrids were clearly differentiated using the melting temperature of the dissociation curves, with derivative peaks at 72 Celsius for An. arabiensis, 75°C for An. gambiae and 86°C for An. coluzzii. The hybrids (An. gambiae / An. coluzzii) showed both peaks. This work is the first to describe a SYBR green real time PCR method for the characterization of An. arabiensis, An. gambiae and An. coluzzii and was purposely designed for basic melt-curve analysis (rather than high-resolution melt-curve) to allow it to be used on a wide range of real-time PCR machines. |
| Databáze: | OpenAIRE |
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