Using DNA Fingerprinting to Identify the Blood-Feeding Patterns of Mosquito Vectors of West Nile Virus Theodore G. Andreadis The Connecticut Agricultural Experiment Station New Haven, CT Introduction • WNV has been detected in 18 different mosquito species in Connecticut • 5 species have been implicated as primary or secondary vectors • Physiologically competent • Frequently infected in nature • Closely associated with virus foci (human cases) Primary Vectors Culex pipiens Culex restuans Culex salinarius Secondary Vectors Aedes vexans Culiseta melanura West Nile Virus Transmission Cycle ? Enzootic Cycle Epidemic ? ? Transmission Wild Bird Reservoirs Mosquito Blood Feeding Study • To examine the feeding patterns of the principal mosquito vectors of WNV in Connecticut Birds Mammals • Identify specific bird and mammal hosts that mosquitoes feed on in nature • Determine the role of these animals in the ecology of WNV in the region Cs. melanura Cx. pipiens Cx. restuans Cx. salinarius Ae. vexans CO2 -baited CDC light trap Gravid trap Mosquito Collection Sites N = 80 Materials and Methods • Abdomens were removed under a dissecting microscope and DNA was isolated • The DNA was amplified by PCR using Avian and Mammalian specific primer pairs to the cytochrome b gene • The PCR amplification products were sequenced and identified to species by comparison to the Genbank® sequence data base Culex Mosquitoes 100 Bird Mammal 80 Bird & Mammal Amphibian 60 1.5% Percent 40 0.5% Human Human 20 0 Culex pipiens Culex restuans Culex salinarius Secondary Mosquito Vectors 100 Bird Mammal 80 Bird & Mammal 60 Percent 40 20 0 Culiseta melanura Secondary Mosquito Vectors 100 Bird Mammal 80 Bird & Mammal 60 Percent 40 20 0 Culiseta melanura Aedes vexans Percentage of Bird-Derived Blood Meals (n = 330) 40 42 different species 30 of birds identified 20 Percent of Total 10 0 American Gray Catbird House Chickadee Mourning American Robin Sparrow Dove Crow Percentage of Bird-Derived Blood Meals (n = 330) 40 • Abundant • Frequently infected 30 Æ • Reservoir competent • Are likely to be a 20 very important source of virus Percent of Total 10 0 American Gray Catbird House Chickadee Mourning American Robin Sparrow Dove Crow Percentage of Bird-Derived Blood Meals (n = 330) 40 • Crows acquire WNV through means 30 other than mosquito bites 20 ? • Crows are not likely to be the primary amplifying hosts Percent of Total 10 0 American Gray Catbird House Chickadee Mourning American Robin Sparrow Dove Crow Percentage of Mammal-Derived Blood Meals (n = 198) 80 70 12 different species of 60 mammals identified 50 40 30 Percent of Total 20 10 0 Deer Horse Cat Raccoon Rabbit Human Percentage of Mammal-Derived Blood Meals (n = 198) 80 • Most abundant large 70 Æ mammal in region 60 • Role in transmission unknown 50 • May divert feeding 40 from horses and humans 30 Percent of Total 20 10 0 Deer Horse Cat Raccoon Rabbit Human Percentage of Mammal-Derived Blood Meals (n = 198) 80 70 • Low level of feeding on humans 60 • Helps to explain 50 limited number of 40 human cases 30 Percent of Total 20 10 0 Deer Horse Cat Raccoon Rabbit Human Northeastern US West Nile Virus Transmission Cycle Overwinter Epidemic Transmission Culex salinarius Culex pipiens Culex pipiens Culex restuans Aedes vexans Culiseta melanura Enzootic June Cycle to Incidental Infections October Virus August to October Wild Bird Reservoir and Amplifying Hosts CAES Mosquito / West Nile Virus Research Group Mosquito Identification & Trapping John Shepard Michael Thomas Blood Meal Analysis Dr. Goudarz Molaei Virus Isolation & Identification Dr. Philip Armstrong Shannon Finan Dr. John Anderson Dr. Charles Vossbrinck.