(a) Nests survey and bird observation in the field
A field survey of nesting birds was conducted over the breeding season from mid-March to late August, 2011–2013, at the Xishuangbanna Tropical Botanical Garden (XTBG; centred at 21°55′N, 101°16′E) in southwest China. Nests were identified to species by direct observation after they were found. Each nest was checked randomly 1–10 times over the breeding period to record whether nestling feces was present. As each check was conducted quietly and quickly (often less than 10 s), and one time a day to decrease disturbance; no nests were abandoned because of our visitation. A similar survey was conducted in XTBG between April-May 2014, in which we recorded not only feces, but also whether adult birds were present in the nest at the time it was checked.
To identify the potential factors that regulated the defecation time under natural conditions, we conducted detailed observations on five nests of the Red-whiskered Bulbul (Pycnonotus jocosus) in the field (mean = 2.4 nestlings/nest, SD = 0.55, n = 5), because this species is abundant and its nests are accessible for observation, often being less than 2 m above the ground. The field observation was conducted using a digital video camera (Sony HDR-XR550), placed at a distance of 0.5-2 m to the nests. Nestlings were marked by the use of non-toxic paints on the top of their heads for individual identification. The first 20 min of observation was excluded from the analysis to permit the parents to return to their normal behaviour. Films were later viewed to document the time of each feeding and defecation, and the status of parent birds (present or absent) when defecation occurred. If only one nestling was fed at a feeding bout, we investigated whether only it defecated. All videotaped individuals fledged.
(b) Laboratory experiment
To investigate the degree of control nestlings have in waiting to defecate after a feeding, we turned to a more controlled laboratory environment, in which the “parents” were people feeding the birds.
We relocated 6 different P. jocosus nests to the laboratory (mean = 2.7 nestlings/nest, SD = 0.52, n = 6), three nests each containing 2 day old nestlings and three nests containing 6 day old nestlings. Each nest was fed for between 1 and 2 min at 1 h, 2 h and 3 h time intervals consecutively in that order, and then this feeding protocol was repeated. Nestlings were fed the same food quantity and items at each feeding; food consisted of primarily mealworms and fruit pellets, comparable to the diet fed to nestlings by parents. We watched the young birds for evidence of undernourishment under these conditions, but their growth appeared similar to the wild offspring of the same age and all these laboratory raised birds were successfully fledged. They were then kept in an aviary for separate experiments.
The data we collected in this experiment included the number of sacs excreted during feeding and non-feeding periods. For the nests containing 2 day old nestlings the experiment lasted 6 days, and for the nests with 6 day old nestlings the experiment lasted 2–3 days. After each feeding visit, we removed all the fecal sacs in the nest to keep it clean. For the 2 day old nestlings, we determined the wet weight of each fecal sac (nearest 0.01 g) at about 10–20 min after they were collected. To increase the sample size for this fecal sac weight data, we moved one additional Red-whiskered Bulbul nest to the lab when the nestlings were two days old (hence the fecal weight data has sample size n = 4 nests).
This research was approved by the Administrative Panel on the Ethics of Animal Experiments of the Xishuangbanna Tropical Botanical Garden, Chinese Academy of Sciences (protocol: XTBG 2011–003), and strictly adhered to the Guideline for the Care and Use of Laboratory Animals in China.
(c) Statistical analysis
The time interval of feedings was calculated as the amount of time between two consecutive feedings of the same individual; the time interval of defecation was also the amount of time between two consecutive defecations for the same nestling. We then asked if there was a relationship between a feeding or defecation interval and the one that followed it. We used a general linear mixed model, with the earlier interval as the fixed factor and nestling (nested within nest) as a random factor. To calculate a pseudo R-squared for these models, we followed Nakagawa and Schielzeth .
To analyze whether in the laboratory experiment treatment affected fecal wet weight, we also applied a general linear mixed model. For this model, treatment was the fixed factor, and nest was the random factor (data was not available per individual nestling).
Generalized linear mixed models were implemented using the lme4 package  in R (R Core Team, version 3.1.2, 2014).