Premise of the study:To study pollination networks in a changing environment, we need accurate, high-throughput methods. Previous studies have shown that more highly resolved networks can be constructed by studying pollen loads taken from bees, relative to field observations. DNA metabarcoding potentially allows for faster and finer-scale taxonomic resolution of pollen compared to traditional approaches (e.g., light microscopy), but has not been applied to pollination networks.Methods:We sampled pollen from 38 bee species collected in Florida from sites differing in forest management. We isolated DNA from pollen mixtures and sequenced rbcL and ITS2 gene regions from all mixtures in a single run on the Illumina MiSeq platform. We identified species from sequence data using comprehensive rbcL and ITS2 databases.Results:We successfully built a proof-of-concept quantitative pollination network using pollen metabarcoding.Discussion:Our work underscores that pollen metabarcoding is not quantitative but that quantitative networks can be constructed based on the number of interacting individuals. Due to the frequency of contamination and false positive reads, isolation and PCR negative controls should be used in every reaction. DNA metabarcoding has advantages in efficiency and resolution over microscopic identification of pollen, and we expect that it will have broad utility for future studies of plant–pollinator interactions.

H I G H L I G H T S• This study documents the increasing importance of on-road emissions of NH 3.• NO x emissions control mechanisms frequently result in NH 3 production and emissions.• NH 3 is an important driver of N deposition in urban-affected areas and near roadways.• NH 4 -N:NO 3 -N ratios in urban deposition are indicative of elevated NH 3 emissions.• On-road NH 3 emissions exceed agricultural emissions where 40% of the U.S. resides. We provide updated spatial distribution and inventory data for on-road NH 3 emissions for the continental United States (U.S.) On-road NH 3 emissions were determined from on-road CO 2 emissions data and empirical NH 3 :CO 2 vehicle emissions ratios. Emissions of NH 3 from on-road sources in urbanized regions are typically 0.1-1.3 t km −2 yr −1 while NH 3 emissions in agricultural regions generally range from 0.4-5.5 t km G R A P H I C A L A B S T R A C T a b s t r a c t a r t i c l e i n f o, with a few hotspots as high as 5.5-11.2 t km −2 yr −1. Counties with higher vehicle NH 3 emissions than from agriculture include 40% of the U.S. population. The amount of wet inorganic N deposition as NH 4 + from the National Atmospheric Deposition Program (NADP) network ranged from 37 to 83% with a mean of 58.7%. Only 4% of the NADP sites across the U.S. had b45% of the N deposition as NH 4 + based on data from 2014 to 2016, illustrating the near-universal elevated proportions of NH 4 + in deposition across the U.S. Case studies of on-road NH 3 emissions in relation to N deposition include four urban sites in Oregon and Washington where the average NH 4 -N:NO 3 -N ratio in bulk deposition was 2.3. At urban sites in the greater Los Angeles Basin, bulk deposition of NH 4 -N and NO 3 -N were equivalent, while NH 4 -N:NO 3 -N in throughfall under shrubs ranged from 0.6 to 1.7. The NH 4 -N:NO 3 -N ratio at 7-10 sites in the Lake Tahoe Basin averaged 1.4 and 1.6 in bulk deposition and throughfall, and deposition of NH 4 -N was strongly correlated with summertime NH 3 concentrations. On-road

Anthropogenic resource supplementation can shape wildlife disease directly by altering the traits and densities of hosts and parasites or indirectly by stimulating prey, competitor or predator species. We first assess the direct epidemiological consequences of supplementation, highlighting the similarities and differences between food provisioning and two widespread forms of nutrient input: agricultural fertilization and aquatic nutrient enrichment. We then review an aquatic disease system and a general model to assess whether predator and competitor species can enhance or overturn the direct effects of enrichment. All forms of supplementation can directly affect epidemics by increasing host population size or altering parasite production within hosts, but food provisioning is most likely to aggregate hosts and increase parasite transmission. However, if predators or competitors increase in response to supplementation, they could alter resource-fuelled outbreaks in focal hosts. We recommend identifying the traits of hosts, parasites or interacting species that best predict epidemiological responses to supplementation and evaluating the relative importance of these direct and indirect mechanisms. Theory and experiments should examine the timing of behavioural, physiological and demographic changes for realistic, variable scenarios of supplementation. A more integrative view of resource supplementation and wildlife disease could yield broadly applicable disease management strategies.This article is part of the theme issue 'Anthropogenic resource subsidies and host-parasite dynamics in wildlife'.

No abstract