I will manipulate the intensity and timing of herbivory on common milkweed at the University of Michigan Biological Station (UMBS) in Pellston, Michigan. Common milkweed does not typically bloom until its second year, so I will obtain mature plants (at least 1 year old) from a commercial grower. Plants will be grown in large pots to simulate a natural setting and not confine the roots. Pots will be placed in a full factorial blocked design containing 15 blocks. Each block will contain six pots of milkweed, each being exposed to one of six treatments: 1) low and early herbivory, 2) low and late herbivory, 3) high and early herbivory, 4) high and late herbivory, 5) no herbivory (control), and 6) natural herbivory. Pots will be placed at least 1 m apart, and blocks will be placed at least 20 m apart to ensure independence and avoid perturbations of pollinators during surveys. Plants in the low herbivory treatments will have 5% of each leaf manually damaged, while those in high herbivory treatments will have 50% of each leaf manually damaged. Herbivory intensity can vary between years and locations; therefore, the chosen percentages are based on past observations of common milkweed at UMBS. Larger plants will by default have more total biomass removed; to account for plant size, plant height and number of leaves will be measured. Early-onset herbivory treatments will begin in early-June to reflect peak damage from early active herbivores, while late-onset herbivory treatments will begin in early-July to reflect peak damage from later occurring herbivores. Leaves will be damaged once on the first day of each treatment by removing leaf tissue using scissors. Plants in all treatments (except natural) will be netted with lightweight material to prevent interference from naturally occurring herbivory. Once blooming begins, only the flowers will be exposed from the netting to allow for pollination to occur.
Pollinators: Since the blooming time of all plants may not be aligned, pollinator surveys will be conducted during the blooming period of individual milkweed plants. Surveys will begin when the first umbel is open and will end when the last umbel begins to senesce. Surveys will also only occur between 10:00 and 16:00, with light winds (<29 km/hr, Beaufort 5), low cloud cover (< 50%), and when air temperature is above 13°C. Plants will be observed for five minutes in random order. During these five minutes, three observers will record the number of each pollinator species or morphotypes (e.g., bees, wasps, butterflies, moths, beetles, flies, and ants) that visit the plant and the duration of time spent on the plant. A visit is characterized by contact between the pollinator's body and the reproductive parts of a flower. Duration will be measured from time of first visit to time of departure from a single plant. Individuals that are not immediately obvious to which group they belong to will be captured with a net, identified through the net, and then released. Possible characteristics for identification include the number and shape of wings, body shape, antennae shape, and elytra. Individuals who cannot be identified in the field will be netted, placed into a killing jar, and identified in the lab.
Flowers traits: On pollinator survey days, floral display size (number of open flowers), number of inflorescences, mean inflorescence size (flowers per umbel), flower area, hood height, nectar volume, and nectar sugar concentration. Individual flower size measurements will be made on five randomly assigned flowers using a caliper. Nectar will be collected from another set of five randomly assigned flowers. Chosen flowers will be covered by lightweight nets the night prior to sampling to prevent pollinators from removing any nectar. Nectar quality will be measured by its sugar concentration in mg/uL, which will be measured using a refractometer. Nectar quantity will be measured by its volume, which will be collected by inserting glass capillary tubes into the flower hoods and measuring the length of the nectar column. Pollen will not be measured as it is inaccessible to pollinators (only pollinia is accessible), making nectar the only available reward. All flower traits will receive additional measurements on non-survey days throughout the blooming period.
Environmental: On the days in which pollinators are observed, temperature, cloud cover, humidity, and wind speed will be measured to rule out any confounding effects. Insect pollinator visitation can vary depending on these variables, they are generally more active when it’s sunny, warm, and with low wind speeds.
Plan B: If our plants are not showing enough progress prior to the damage, our plan B would be to use the populations of naturally growing milkweed in the ballfield. In this case, we would need to choose stems with no weevil damage before netting. Another option is to put mesh sleeves into the ground to prevent weevil damage. We would replace the block system with genets, and individual plants would be renets.