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DNA barcoding: complementing morphological identification of mosquito species in Singapore.

ABSTRACT: Taxonomy that utilizes morphological characteristics has been the gold standard method to identify mosquito species. However, morphological identification is challenging when the expertise is limited and external characters are damaged because of improper specimen handling. Therefore, we explored the applicability of mitochondrial cytochrome C oxidase subunit 1 (COI) gene-based DNA barcoding as an alternative tool to identify mosquito species. In the present study, we compared the morphological identification of mosquito specimens with their differentiation based on COI barcode, in order to establish a more reliable identification system for mosquito species found in Singapore.We analysed 128 adult mosquito specimens, belonging to 45 species of 13 genera. Phylogenetic trees were constructed for Aedes, Anopheles, Culex and other genera of mosquitoes and the distinctive clustering of different species was compared with their taxonomic identity.The COI-based DNA barcoding achieved a 100% success rate in identifying the mosquito species. We also report COI barcode sequences of 16 mosquito species which were not available previously in sequence databases.Our study utilised for the first time DNA barcoding to identify mosquito species in Singapore. COI-based DNA barcoding is a useful tool to complement taxonomy-based identification of mosquito species.

SUBMITTER: Chan A

PROVIDER: S-EPMC4282734 | biostudies-literature | 2014

REPOSITORIES: biostudies-literature

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DNA barcoding: complementing morphological identification of mosquito species in Singapore.

Chan Abigail A Chiang Lee-Pei LP Hapuarachchi Hapuarachchige C HC Tan Cheong-Huat CH Pang Sook-Cheng SC Lee Ruth R Lee Kim-Sung KS Ng Lee-Ching LC Lam-Phua Sai-Gek SG

Parasites & vectors 20141212

<h4>Background</h4>Taxonomy that utilizes morphological characteristics has been the gold standard method to identify mosquito species. However, morphological identification is challenging when the expertise is limited and external characters are damaged because of improper specimen handling. Therefore, we explored the applicability of mitochondrial cytochrome C oxidase subunit 1 (COI) gene-based DNA barcoding as an alternative tool to identify mosquito species. In the present study, we compared ...[more]

PMID: 25498759

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Project description:BackgroundIdentification of mosquitoes greatly relies on morphological specification. Since some species cannot be distinguished reliably by morphological methods, it is important to incorporate molecular techniques into the diagnostic pipeline. DNA barcoding using Sanger sequencing is currently widely used for identification of mosquito species. However, this method does not allow detection of multiple species in one sample, which would be important when analysing mosquito eggs. Detection of container breeding Aedes is typically performed by collecting eggs using ovitraps. These traps consist of a black container filled with water and a wooden spatula inserted for oviposition support. Aedes mosquitoes of different species might lay single or multiple eggs on the spatula. In contrast to Sanger sequencing of specific polymerase chain reaction (PCR) products, multiplex PCR protocols targeting specific species of interest can be of advantage for detection of multiple species in the same sample.MethodsFor this purpose, we adapted a previously published PCR protocol for simultaneous detection of four different Aedes species that are relevant for Austrian monitoring programmes, as they can be found in ovitraps: Aedes albopictus, Aedes japonicus, Aedes koreicus, and Aedes geniculatus. For evaluation of the multiplex PCR protocol, we analysed 2271 ovitrap mosquito samples from the years 2021 and 2022, which were collected within the scope of an Austrian nationwide monitoring programme. We compared the results of the multiplex PCR to the results of DNA barcoding.ResultsOf 2271 samples, the multiplex PCR could identify 1990 samples, while species determination using DNA barcoding of the mitochondrial cytochrome c oxidase subunit I gene was possible in 1722 samples. The multiplex PCR showed a mixture of different species in 47 samples, which could not be detected with DNA barcoding.ConclusionsIn conclusion, identification of Aedes species in ovitrap samples was more successful when using the multiplex PCR protocol as opposed to the DNA barcoding protocol. Additionally, the multiplex PCR allowed us to detect multiple species in the same sample, while those species might have been missed when using DNA barcoding with Sanger sequencing alone. Therefore, we propose that the multiplex PCR protocol is highly suitable and of great advantage when analysing mosquito eggs from ovitraps.

Dataset Information

DNA barcoding: complementing morphological identification of mosquito species in Singapore.

Publications

DNA barcoding: complementing morphological identification of mosquito species in Singapore.

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