Wild duck species sampled
Surveillance of wild ducks in Alberta, Canada was initiated in 1976 in conjunction with the Canadian Wildlife Service (Prairie and Northern Region) from late July through early September. This period of months is the time when the breeding season is completed, many of the adult ducks have molted, and the birds are staging before they migrate south to their winter habitat. During the 26 years of the study 17592 cloacal swabs were collected from wild ducks.

The major species sampled were:

• mallard ( Anas platyrhynchos ) 64.4%
• northern pintail ( Anas acuta ) 17.7%
• blue-winged teal ( Anas discors ) 10.7%
• redhead ( Aythya Americana ) 2.3%
• American green-winged teal ( Anas crecca ) 1.1%
• American wigeon ( Anas Americana ) 1.1%.
• The remaining 2.7% of the sampled birds was represented by:
• black duck ( Anas rubripes )
• bufflehead ( Bucephala albeola )
• Canada goose ( Branta canadensis )
• canvasback ( Aythya valisineria )
• cinnamon teal ( Anas cyanoptera )
• common goldeneye ( Bucephala clangula )
• coot ( Fulica americana )
• gadwall ( Anas strepera )
• lesser scaup ( Aythya affinis )
• ruddy duck ( Oxyura jamaicensis )
• northern shoveler ( Anas clypeata ).
  

Virus isolation and serological identification of subtype
The samples were thawed, and 0.3 mL of fluid from the vial was mixed with 0.3 mL PBS containing high concentrations of antibiotics (penicillin G, 4000 U/mL; streptomycin sulfate, 800 U/mL; polymyxin B, 400 U/mL; and gentamycin sulfate, 0.1 mg/mL). For each sample, the allantoic cavities of three 11-day-old embryonated chicken eggs were injected with 0.2 mL of inoculum. The eggs were incubated at 35 °C for 48 to 72 h, and the embryos examined for viability. The eggs then were chilled at 4 o C for 12 to 16 h (overnight). The allantoic fluid of each egg was tested for the presence of agglutinating virus by the hemagglutination assay. The positive samples were checked for bacterial contamination by streaking blood-agar plates and then incubating the plates at 37 o C for 48 h. Bacterial contamination was removed by passing the allantoic fluid through a 0.2- m M syringe filter.

To determine the hemagglutinin (HA) subtype, samples that were positive for hemagglutination were tested in hemagglutination inhibition (HAI) assays, and the neuraminidase (NA) subtype was determined by the neuraminidase inhibition (NI) assay. Hemagglutinating agents were identified on the basis of their antigenic characteristics (inhibition) by using monospecific antisera prepared against the surface antigens of reference influenza A viruses. When monospecific serum was not available, chicken or rabbit antiserum prepared against whole-virus antigen was used.