Journal of the American Mosquito Control Association (2007) 23, 488-491

Ilia Rochlin, Michael P. Santoriello, Richard T. Mayer and Scott R. Campbell (2007)
Improved high-throughput method for molecular identification of Culex mosquitoes
Journal of the American Mosquito Control Association 23 (4), 488-491
Abstract: Culex pipiens, Cx. restuans, and Cx. salinarius play important and most likely different roles in transmission of West Nile virus (WNV) in the northeastern United States. While Cx. pipiens and Cx. restuans are considered the main enzootic vectors of WNV, Cx. salinarius may be involved in epizootic cycles due to its broader host preferences. Accurate morphological identification of field-collected Culex specimens may be difficult, and therefore the New York State Department of Health arbovirus surveillance program allows combined Cx. pipiens and Cx. restuans pools to be tested for WNV. We have developed a modified and improved DNA isolation protocol using proteinase K digestion without traditional mosquito trituration and nucleic acid extraction to permit high-throughput screening of a large number of Culex specimens for species identification using polymerase chain reaction (PCR). This method utilizes a 96-well-plate format and a novel 1-step crude extraction procedure using proteinase K to obtain genomic DNA template from 1 mosquito leg in sufficient quantity for at least 2 standard 50-µl PCR reactions. Proteinase K digestion of legs from individual Culex mosquitoes was performed and used for PCR amplification with previously described species-specific ribosomal DNA primers. Using these rDNA primers, our modified proteinase K method successfully identified 91% to 100% of the Culex samples.
(The abstract is excluded from the Creative Commons licence and has been copied with permission by the publisher.)
Link to article at publishers website

Research topic(s) for pests/diseases/weeds:
identification/taxonomy
surveys/sampling/distribution

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Pest and/or beneficial records:

Beneficial	Pest/Disease/Weed	Crop/Product	Country	Quarant.
	Culex pipiens		U.S.A. (NE)
	Culex salinarius		U.S.A. (NE)
	Culex restuans		U.S.A. (NE)