Experimental Design and Field Methodology
At each field site, we placed a pair of B. vosnesenskii colonies
on a given date. Within the pair, we randomly assigned colonies to one
of two food treatments: early supplementation (during the first 20 days
in the field, early-pulse) or late supplementation (at 21-40 days in the
field, late-pulse). In each treatment, food pulses were intended to
represent elevated resources occurring earlier or later during the
growth phase of colony dynamics (based on our past experience withB. vosnesenskii, cf. Crone & Williams 2016; Kerr et al.2019; Malfi et al. 2019). Colony weight at field placement did
not differ by experimental food treatment (F1,26 = 2.04,P = 0.17).
During pulses, a colony received (1) ad libitum access to
artificial nectar (BioGluc®, BioBest Canada Ltd.), which was delivered
directly to the colony via a reservoir with an access point located
inside of the colony box (Fig S1), and (2) pollen in the form fresh
frozen honey bee pollen mixed with diluted artificial nectar at a
~1:1 mass ratio, and encased in USDA certified organic
beeswax (Koster-Keunan Inc., CT, USA), delivered to the colony on the
first and 10th day of the 20-day pulse. The total
weight of pollen provided to a colony was proportional to the colony
weight at the time of each feeding in order to provide colonies 50% of
their estimated needs for a 10-day interval (see Supporting
Information, Appendix 3 ). We used this limit because superabundance of
pollen in the nest can reduce foraging activity (Pelletier & McNeil
2003). To determine whether and how foraging was affected by
supplementation, we recorded the number of return trips per hour for
both colonies during site visits throughout the experiment and analyzed
this foraging activity information in relation to food treatment (seeSupporting Information, Appendix 4 ). At the end of the 20-day
pulse, access to food supplements was terminated.
We weighed colonies in the field every 10 days in order to track their
growth across the season. We assumed that a colony’s peak weight would
be associated with its switch to reproduction (Crone & Williams 2016)
and pulled them from the field once they had declined in weight for two
consecutive visits. This ensured that brood structure of the nest would
be intact, allowing for final counting of reproductive output. Declining
colonies are commonly invaded by insects and fungi that quickly destroy
the nest structure. Upon removal from the field, colonies were frozen
(-20º C) until they could be dissected in the laboratory. Gyne
production was determined by counting the number of queen brood cells
present during dissections of the remnant brood structure. Queen cells
can be reliably distinguished from worker/male cells based on their
larger size (Williams et al. 2012). At the time of dissection,
the majority of gynes had already eclosed; for the few colonies where
some gynes were still in cocoons, both eclosed and uneclosed gynes were
tabulated in the total gyne count.