Utility of carbon and nitrogen stable isotopes for inferring wild bee (Hymenoptera: Apoidea) use of adjacent foraging habitats.

Isotope analysis has proven useful for understanding diets of animals that are difficult to track for extended periods. Bees are small yet highly mobile and often forage from multiple habitats. However, current methods of assessing diet are limited in scope. Efficient methods of tracking bee diets that integrate across life stages, distinguish habitat use, and are sensitive to taxonomic differences will inform conservation strategies. We evaluated the utility of stable isotope analysis for estimating contributions of adjacent habitats to bees' diets. We also investigated taxonomic variation in bee and flower isotope composition. We measured natural abundance of carbon and nitrogen stable isotopes in two body regions from three wild bee genera, as well as in 25 species of flowers that likely comprised their diets. Bee ∂13C and ∂15N varied with habitat and taxonomic groups (conflated with month), but did not match spatial or seasonal trends in their food plants. Flower ∂13C was lowest in the forest and in April-June, as expected if driven by water availability. However, bee ∂13C was elevated in the spring, likely from overwintering nutritional stress or unpredictable food availability. Bumble bees (Bombus) were enriched in ∂15N compared to others, possibly reflecting differences in larval feeding. Bee diet mixing models had high variation and should be interpreted with caution. Models estimated similar habitat contributions to diets of spring Andrena and overwintered Bombus queens. Summer Bombus queens and workers were indistinguishable. Sweat bees (Halictus) were estimated to use comparatively more field flowers than others. Overall, taxon more strongly influenced isotope composition than either foraging habitat or month, likely because of associated differences in sociality and timing of annual activity. Future studies seeking to reveal bee diets by isotope analysis may gain better resolution in more isotopically distinct habitats, in conjunction with controlled feeding or isotope labeling experiments.

Ten-year trends reveal declining quality of seeded pollinator habitat on reclaimed mines regardless of seed mix diversity.

Plant-pollinator interactions represent a crucial ecosystem function threatened by anthropogenic landscape changes. Disturbances that reduce plant diversity are associated with floral resource and pollinator declines. Establishing wildflower plantings is a major conservation strategy targeting pollinators, the success of which depends on long-term persistence of seeded floral communities. However, most pollinator-oriented seeding projects are monitored for a few years, making it difficult to evaluate the longevity of such interventions. Selecting plant species to provide pollinators diverse arrays of floral resources throughout their activity season is often limited by budgetary constraints and other conservation priorities. To evaluate the long-term persistence of prairie vegetation seeded to support pollinators, we sowed wildflower seed mixes into plots on a degraded reclaimed strip-mine landscape in central Ohio, USA. We examined how pollinator habitat quality, measured as floral abundance and diversity, changed over 10 years (2009-2019) in the absence of management, over the course of the blooming season within each year, and across three seed mixes containing different numbers and combinations of flowering plant species. Seeded species floral abundance declined by more than 75% over the study, with the largest decline occurring between the fifth and seventh summers. Native and non-native adventive flowering plants quickly colonized the plots and represented >50% of floral community abundances on average. Floral richness remained relatively constant throughout the study, with a small peak one year after plot establishment. Plots seeded with High-Diversity Mixes averaged two or three more species per plot compared with a Low-Diversity Mix, despite having been seeded with twice as many plant species. Within years, the abundance and diversity of seeded species were lowest early in the blooming season and increased monotonically from June to August. Adventive species exhibited the opposite trend, such that complementary abundance patterns of seeded and adventive species blooms resulted in a relatively constant floral abundance across the growing season. Seeded plant communities followed classic successional patterns in which annual species quickly established and flowered but were replaced by perennial species after the first few summers. Long-term data on establishment and persistence of flower species can guide species selection for future-oriented pollinator habitat restorations.

Preference of Peponapis pruinosa (Hymenoptera: Apoidea) for Tilled Soils Regardless of Soil Management System.

Concerns about global pollinator declines have placed a growing focus on understanding the impact of agriculture practices on valuable native pollinators in these systems. Cultivation practices such as tillage disturb agroecosystems and can have negative impacts on ground-nesting pollinators. The squash bee, Peponapis pruinosa (Say), is a ground-nesting specialist pollinator of Cucurbita (Cucurbitaceae) crops (i.e., pumpkins and squash) that often nests in agricultural fields and thus may be vulnerable to these practices. We investigated the impact of tillage on nesting behavior of P. pruinosa in plasticulture and strip-tilled squash systems. We used choice experiments to test nesting substrate preference and nesting success of caged P. pruinosa in two soil tillage systems: strip tillage and plasticulture. The strip tillage system comprised two tillage zones (strip-tilled row with no-till edges), and the plasticulture system comprised two tillage zones (plastic bed and conventional tillage edge). The results of our study indicate that P. pruinosa nesting density did not significantly differ between the strip tillage and plasticulture systems. Within each system, P. pruinosa preferred excavating nests in the most disturbed soil zones (strip-tilled row and conventionally tilled edge). In the strip tillage system, the strip-tilled row had significantly more nests than the no-till edge. Results of these studies suggest that soil tillage practices can influence P. pruinosa nesting choice and production practices should be considered when developing a pollinator protection plan.

Invasion of a dominant floral resource: effects on the floral community and pollination of native plants.

Through competition for pollinators, invasive plants may suppress native flora. Community-level studies provide an integrative assessment of invasion impacts and insights into factors that influence the vulnerability of different native species. We investigated effects of the nonnative herb Lythrum salicaria on pollination of native species in 14 fens of the eastern United States. We compared visitors per flower for 122 native plant species in invaded and uninvaded fens and incorporated a landscape-scale experiment, removing L. salicaria flowers from three of the invaded fens. Total flower densities were more than three times higher in invaded than uninvaded or removal sites when L. salicaria was blooming. Despite an increase in number of visitors with number of flowers per area, visitors per native flower declined with increasing numbers of flowers. Therefore, L. salicaria invasion depressed visitation to native flowers. In removal sites, visitation to native flowers was similar to uninvaded sites, confirming the observational results and also suggesting that invasion had not generated a persistent build-up of visitor populations. To study species-level impacts, we examined effects of invasion on visitors per flower for the 36 plant species flowering in both invaded and uninvaded fens. On average, the effect of invasion represented about a 20% reduction in visits per flower. We measured the influence of plant traits on vulnerability to L. salicaria invasion using meta-analysis. Bilaterally symmetrical flowers experienced stronger impacts on visitation, and similarity in flower color to L. salicaria weakly intensified competition with the invader for visitors. Finally, we assessed the reproductive consequences of competition with the invader in a dominant flowering shrub, Dasiphora fruticosa. Despite the negative effect of invasion on pollinator visitation in this species, pollen limitation of seed production was not stronger in invaded than in uninvaded sites, suggesting little impact of competition for pollinators on its population demography. Negative effects on pollination of native plants by this copiously flowering invader appeared to be mediated by increases in total flower density that were not matched by increases in pollinator density. The strength of impact was modulated across native species by their floral traits and reproductive ecology.

Demographic consequences of greater clonal than sexual reproduction in Dicentra canadensis.

Clonality is a widespread life history trait in flowering plants that may be essential for population persistence, especially in environments where sexual reproduction is unpredictable. Frequent clonal reproduction, however, could hinder sexual reproduction by spatially aggregating ramets that compete with seedlings and reduce inter-genet pollination. Nevertheless, the role of clonality in relation to variable sexual reproduction in population dynamics is often overlooked. We combined population matrix models and pollination experiments to compare the demographic contributions of clonal and sexual reproduction in three Dicentra canadensis populations, one in a well-forested landscape and two in isolated forest remnants. We constructed stage-based transition matrices from 3 years of census data to evaluate annual population growth rates, λ. We used loop analysis to evaluate the relative contribution of different reproductive pathways to λ. Despite strong temporal and spatial variation in seed set, populations generally showed stable growth rates. Although we detected some pollen limitation of seed set, manipulative pollination treatments did not affect population growth rates. Clonal reproduction contributed significantly more than sexual reproduction to population growth in the forest remnants. Only at the well-forested site did sexual reproduction contribute as much as clonal reproduction to population growth. Flowering plants were more likely to transition to a smaller size class with reduced reproductive potential in the following year than similarly sized nonflowering plants, suggesting energy trade-offs between sexual and clonal reproduction at the individual level. Seed production had negligible effects on growth and tuber production of individual plants. Our results demonstrate that clonal reproduction is vital for population persistence in a system where sexual reproduction is unpredictable. The bias toward clonality may be driven by low fitness returns for resource investment in sexual reproduction at the individual level. However, chronic failure in sexual reproduction may exacerbate the imbalance between sexual and clonal reproduction and eventually lead to irreversible loss of sex in the population.

Rank-based characterization of pollen assemblages collected by honey bees using a multi-locus metabarcoding approach.

PREMISE OF THE STUDY: Difficulties inherent in microscopic pollen identification have resulted in limited implementation for large-scale studies. Metabarcoding, a relatively novel approach, could make pollen analysis less onerous; however, improved understanding of the quantitative capacity of various plant metabarcode regions and primer sets is needed to ensure that such applications are accurate and precise. METHODS AND RESULTS: We applied metabarcoding, targeting the ITS2, matK, and rbcL loci, to characterize six samples of pollen collected by honey bees, Apis mellifera. Additionally, samples were analyzed by light microscopy. We found significant rank-based associations between the relative abundance of pollen types within our samples as inferred by the two methods. CONCLUSIONS: Our findings suggest metabarcoding data from plastid loci, as opposed to the ribosomal locus, are more reliable for quantitative characterization of pollen assemblages. Furthermore, multilocus metabarcoding of pollen may be more reliable than single-locus analyses, underscoring the need for discovering novel barcodes and barcode combinations optimized for molecular palynology.

Application of ITS2 metabarcoding to determine the provenance of pollen collected by honey bees in an agroecosystem.

PREMISE OF THE STUDY: Melissopalynology, the identification of bee-collected pollen, provides insight into the flowers exploited by foraging bees. Information provided by melissopalynology could guide floral enrichment efforts aimed at supporting pollinators, but it has rarely been used because traditional methods of pollen identification are laborious and require expert knowledge. We approach melissopalynology in a novel way, employing a molecular method to study the pollen foraging of honey bees (Apis mellifera) in a landscape dominated by field crops, and compare these results to those obtained by microscopic melissopalynology. • METHODS: Pollen was collected from honey bee colonies in Madison County, Ohio, USA, during a two-week period in midspring and identified using microscopic methods and ITS2 metabarcoding. • RESULTS: Metabarcoding identified 19 plant families and exhibited sensitivity for identifying the taxa present in large and diverse pollen samples relative to microscopy, which identified eight families. The bulk of pollen collected by honey bees was from trees (Sapindaceae, Oleaceae, and Rosaceae), although dandelion (Taraxacum officinale) and mustard (Brassicaceae) pollen were also abundant. • DISCUSSION: For quantitative analysis of pollen, using both metabarcoding and microscopic identification is superior to either individual method. For qualitative analysis, ITS2 metabarcoding is superior, providing heightened sensitivity and genus-level resolution.

The role of resources and risks in regulating wild bee populations.

Recent declines of bee species have led to great interest in preserving and promoting bee populations for agricultural and wild plant pollination. Many correlational studies have examined the indirect effects of factors such as landscape context and land management practices and found great variation in bee response. We focus here on the evidence for effects of direct factors (i.e., food resources, nesting resources, and incidental risks) regulating bee populations and then interpret varied responses to indirect factors through their species-specific and habitat-specific effects on direct factors. We find strong evidence for food resource availability regulating bee populations, but little clear evidence that other direct factors are commonly limiting. We recommend manipulative experiments to illuminate the effects of these different factors. We contend that much of the variation in impact from indirect factors, such as grazing, can be explained by the relationships between indirect factors and floral resource availability based on environmental circumstances.

Population size and relatedness affect fitness of a self-incompatible invasive plant.

One of the lingering paradoxes in invasion biology is how founder populations of an introduced species are able to overcome the limitations of small size and, in a "reversal of fortune," proliferate in a new habitat. The transition from colonist to invader is especially enigmatic for self-incompatible species, which must find a mate to reproduce. In small populations, the inability to find a mate can result in the Allee effect, a positive relationship between individual fitness and population size or density. Theoretically, the Allee effect should be common in founder populations of self-incompatible colonizing species and may account for the high rate of failed introductions, but little supporting evidence exists. We created a field experiment to test whether the Allee effect affects the maternal fitness of a self-incompatible invasive species, wild radish (Raphanus sativus). We created populations of varying size and relatedness. We measured maternal fitness in terms of both fruit set per flower and seed number per fruit. We found that both population size and the level of genetic relatedness among individuals influence maternal reproductive success. Our results explicitly define an ecological genetic obstacle faced by populations of an exotic species on its way to becoming invasive. Such a mechanistic understanding of the invasions of species that require a mate can and should be exploited for both controlling current outbreaks and reducing their frequency in the future.

Food availability affects Osmia pumila (Hymenoptera: Megachilidae) foraging, reproduction, and brood parasitism.

Food limitation can reduce reproductive success directly, as well as indirectly, if foraging imposes a risk of predation or parasitism. The solitary bee Osmia pumila suffers brood parasitism by the cleptoparasitic wasp Sapyga centrata, which enters the host nest to oviposit while the female bee is away. I studied foraging and reproduction of O. pumila nesting within cages stocked with rich or sparse floral resources, and the presence or absence of S. centrata to test (1) the response of nesting female O. pumila to food shortages, (2) the response of nesting female O. pumila to the presence of parasites, and (3) whether brood produced under scarce resources are more likely to be parasitized by S. centrata. The rate of brood cell production was significantly lower in cages with sparse floral resources, although females in sparse cages did not produce significantly fewer brood cells overall. Sapyga centrata did not influence the rate of brood cell production, but females exposed to the cleptoparasites had marginally significantly lower reproductive output. Nests in parasite cages had significantly fewer brood cells than those in parasite free cages. The mean duration of foraging bouts made by female O. pumila in sparse cages was not significantly longer than that in rich cages. O. pumila spent less time in the nest between pollen and nectar foraging bouts in sparse cages with S. centrata than those in other cages suggesting that these individuals made more frequent food foraging trips. Despite the weak effects of parasites and bloom density on foraging behavior, O. pumila brood cells experienced a 5-fold higher probability of parasitism by S. centrata in cages with sparse bloom than in those with rich bloom [corrected]. These results support the hypothesis that indirect effects of food scarcity increase O. pumila susceptibility to brood parasitism, although the exact mechanism is not entirely clear yet.