The fixed factors age and sex were merely included in the models to conduct the interactions. of the ongoing national wild bird avian influenza computer virus (AIV) surveillance program (dd 2014-09-20), executed by the department of Viroscience of Erasmus MC, where mallards, free-living and hunted in the near surrounding, were sampled for LPAIV from 2005 onwards. Sampling During the LPAIV epizootic (i.e. from August until December 2010) studied here, the duck decoy was frequented, on average, seven occasions per month capturing approximately 11 birds per visit. Each captured mallard was marked using a metal ring with an unique code, aged (juvenile: 1 year, adult: 1 year) and sexed based on plumage characteristics . For computer virus detection, Rabbit polyclonal to AGO2 cloacal and oropharyngeal samples were collected using sterile cotton swabs as LPAIV may replicate in both the intestinal and respiratory tract of wild birds . Swabs EC 144 were stored individually in virus transport medium (Hank’s balanced salt answer with supplements; ) at 4C, and transported to the laboratory for analysis within seven days of collection. For detection of antibodies to AIV, blood samples ( 1 ml, 2% of the circulating blood volume) were collected from your brachial vein, which were allowed to clot for approximately 6 h before centrifugation to separate serum from reddish blood cells . Serum samples were stored at ?20C until analysis. To determine a bird’s migratory strategy using stable hydrogen isotope analysis, the tip (1C2 cm) of the first main feather of the right wing was collected and stored in a sealed bag at room heat. Of recaptured birds, both swabs and a blood sample were collected. Migratory strategy In the study of van Dijk et al. , the origin (and hence, migratory strategy) of mallards sampled during the 2010 LPAIV epizootic was decided using stable hydrogen isotope analysis in feathers. Stable isotope signatures in feathers reflect those of local food webs . During the period of growth (i.e. moult), local precipitation is incorporated into these feathers , causing the stable hydrogen isotope (2H) ratio in feathers to be correlated with 2H of local precipitation . Across Europe, a gradient of 2H in feathers is found in mallards . Based on feather 2H and additional criteria, van Dijk et al.  classified mallards as resident, local migrant (i.e. short distance) and distant migrant (i.e. long distance). A resident bird had produced its feathers near the duck decoy (was captured during moult) and was recaptured multiple occasions either before or during the LPAIV epizootic. A local and distant migratory bird was seen and sampled once, i.e. only during the LPAIV epizootic and was not captured within one year before this epizootic. Based on feather 2H values of local (?103.5 to ?72.6) and distant migrants (?164.5 to ?103.7) and using a Western feather 2H isoscape of mallards , local migrants originated roughly from central Europe and distant migrants roughly from north-eastern Europe. We used comparable criteria to assess the migratory strategy of mallards caught during the H3 LPAIV epizootic. For 149 individual birds in this study we were unable to assign them to either the resident or migratory populace and these were excluded from analyses, except the genetic analysis. For full details on the stable hydrogen isotope analysis, see van Dijk et al. . In short, feathers were washed and air-dried overnight. Feather samples were placed into silver capsules, stored in 96 well trays and shipped to the Colorado Plateau Stable Isotope Laboratory (Northern Arizona University or college, Flagstaff, EC 144 USA). Stable hydrogen isotope analyses were performed on a Delta Plus XL EC 144 isotope ratio mass spectrometer equipped with a 1400 C TC/EA pyrolysis furnace. Feather 2H values are reported in models per mil () relative to the Vienna Standard Mean Ocean Water-Standard Light Antarctic Precipitation (VSMOW-SLAP) standard scale. Virus.