Analyzing Mosquito (Diptera: Culicidae) Diversity in Pakistan by DNA Barcoding

Abstract

BackgroundAlthough they are important disease vectors mosquito biodiversity in Pakistan is poorly known. Recent epidemics of dengue fever have revealed the need for more detailed understanding of the diversity and distributions of mosquito species in this region. DNA barcoding improves the accuracy of mosquito inventories because morphological differences between many species are subtle, leading to misidentifications.Methodology/Principal FindingsSequence variation in the barcode region of the mitochondrial COI gene was used to identify mosquito species, reveal genetic diversity, and map the distribution of the dengue-vector species in Pakistan. Analysis of 1684 mosquitoes from 491 sites in Punjab and Khyber Pakhtunkhwa during 2010–2013 revealed 32 species with the assemblage dominated by Culex quinquefasciatus (61% of the collection). The genus Aedes (Stegomyia) comprised 15% of the specimens, and was represented by six taxa with the two dengue vector species, Ae. albopictus and Ae. aegypti, dominant and broadly distributed. Anopheles made up another 6% of the catch with An. subpictus dominating. Barcode sequence divergence in conspecific specimens ranged from 0–2.4%, while congeneric species showed from 2.3–17.8% divergence. A global haplotype analysis of disease-vectors showed the presence of multiple haplotypes, although a single haplotype of each dengue-vector species was dominant in most countries. Geographic distribution of Ae. aegypti and Ae. albopictus showed the later species was dominant and found in both rural and urban environments.ConclusionsAs the first DNA-based analysis of mosquitoes in Pakistan, this study has begun the construction of a barcode reference library for the mosquitoes of this region. Levels of genetic diversity varied among species. Because of its capacity to differentiate species, even those with subtle morphological differences, DNA barcoding aids accurate tracking of vector populations.

DNA barcoding mosquitoes: advice for potential prospectors

“…Indeed, morphological and molecular comparisons of the COI sequence do appear congruent in studies across genus levels, as attested by morphological species' studies on mosquito diversity from Argentina (Laurito et al 2013), Australia (Batovska et al 2016), China (Wang et al 2012), India (Kumar et al 2007), Singapore (Chan et al 2014), Italy (Talbalaghi andShaikevich, 2011), Iran (Azari-Hamidian et al 2009) and Pakistan (Ashfaq et al 2014). This latter study from Pakistan reveals intraspecific divergences at a maximum of 2.4% from over 1600 individuals from 24 taxa -a result in line with the original Canadian work.…”

Mosquito species (Diptera, Culicidae) in three ecosystems from the Colombian Andes: identification through DNA barcoding and adult morphology

“…In mosquitoes, the effectiveness of the COI barcode marker for specimen identification has been tested in surveys of Canada (Cywinska et al 2006), India (Kumar et al 2007), the Iranian islands in the Persian Gulf (Azari-Hamidian et al 2010), China (Wang et al 2012), Amazonian Ecuador (Linton et al 2013), Pakistan (Ashfaq et al 2014), Singapore (Chan et al 2014), and Belgium (Versteirt et al 2015). These studies show correspondence between morphological species and DNA barcode clusters, but also point out the failure of the methodology to distinguish between very similar or cryptic species.…”

Composition and Genetic Diversity of Mosquitoes (Diptera: Culicidae) on Islands and Mainland Shores of Kenya’s Lakes Victoria and Baringo

“…In other parts of the world, mosquitoes have been successfully differentiated using the mitochondrial cytochrome c oxidase 1 (COI) gene, also known as the DNA barcode gene (Hebert et al. 2003, Ashfaq et al. 2014), and the ribosomal internal spacer region (Sum et al.…”