- INTRODUCTION
- MATERIALS AND METHODS
- Mosquito material
- Nucleic acid isolation
- ITS1 PCR amplification and sequencing
- Real-time PCR primer and probe design
- Real-time PCR amplifications
- Synthetic primer and probe controls
- Validation of real-time PCR assays
- Assessment of the real-time PCR assays using field collected samples
- Evaluation of FTA cards and filter paper for sample preservation
- RESULTS
- PCR amplification and sequencing of ITS1
- Synthetic control development
- Sensitivity of real-time PCR assays
- Specificity of real-time PCR assays
- Identification of different life stages and mixed pools
- Assessment of real-time PCR assays using field collected samples
- Evaluation of FTA cards for sample preservation
- DISCUSSION
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Rapid Identification of Aedes albopictus, Aedes scutellaris, and Aedes aegypti Life Stages Using Real-time Polymerase Chain Reaction Assays
In 2005, a widespread infestation of Aedes albopictus was discovered in the Torres Strait, the region between northern Australia and New Guinea. To contain this species, an eradication program was implemented in 2006. However, the progress of this program is impeded by the difficulty of morphologically separating Ae. albopictus larvae from the endemic species Aedes scutellaris. In this study, three real-time TaqMan polymerase chain reaction assays that target the ribosomal internal transcribed spacer 1 region were developed to rapidly identify Aedes aegypti, Ae. albopictus, and Ae. scutellaris from northern Australia. Individual eggs, larvae, pupae, and adults, as well as the species composition of mixed pools were accurately identified. The assay method was validated using 703 field-collected specimens from the Torres Strait.