Bee monitoring by community scientists: comparing a collections-based program with iNaturalist.

Bee monitoring, or widespread efforts to document bee community biodiversity, can involve data collection using lethal (specimen collections) or non-lethal methods (observations, photographs). Additionally, data can be collected by professional scientists or by volunteer participants from the general public. Collection-based methods presumably produce more reliable data with fewer biases against certain taxa, while photography-based approaches, such as data collected from public natural history platforms like iNaturalist, can involve more people and cover a broader geographic area. Few efforts have been made to quantify the pros and cons of these different approaches. We established a community science monitoring program to assess bee biodiversity across the state of Pennsylvania (USA) using specimen collections with nets, blue vane traps, and bowl traps. We recruited 26 participants, mostly Master Gardeners, from across the state to sample bees after receiving extensive training on bee monitoring topics and methods. The specimens they collected were identified to species, stored in museum collections, and the data added to public databases. Then, we compared the results from our collections to research-grade observations from iNaturalist during the same time period (2021 and 2022). At state and county levels, we found collections data documented over twice as much biodiversity and novel baseline natural history data (state and county records) than data from iNaturalist. iNaturalist data showed strong biases toward large-bodied and non-native species. This study demonstrates the value of highly trained community scientists for collections-based research that aims to document patterns of bee biodiversity over space and time.

Fungicide ingestion reduces net energy gain and microbiome diversity of the solitary mason bee.

Fungicides are frequently used during tree fruit bloom and can threaten insect pollinators. However, little is known about how non-honey bee pollinators such as the solitary bee, Osmia cornifrons, respond to contact and systemic fungicides commonly used in apple production during bloom. This knowledge gap limits regulatory decisions that determine safe concentrations and timing for fungicide spraying. We evaluated the effects of two contact fungicides (captan and mancozeb) and four translaminar/plant systemic fungicides (cyprodinil, myclobutanil, penthiopyrad, and trifloxystrobin) on larval weight gain, survival, sex ratio, and bacterial diversity. This assessment was carried out using chronic oral ingestion bioassays where pollen provisions were treated with three doses based on the currently recommended field use dose (1X), half dose (0.5X), and low dose (0.1X). Mancozeb and penthiopyrad significantly reduced larval weight and survival at all doses. We then sequenced the 16S gene to characterize the larvae bacteriome of mancozeb, the fungicide that caused the highest mortality. We found that larvae fed on mancozeb-treated pollen carried significantly lower bacterial diversity and abundance. Our laboratory results suggest that some of these fungicides can be particularly harmful to the health of O. cornifrons when sprayed during bloom. This information is relevant for future management decisions about the sustainable use of fruit tree crop protection products and informing regulatory processes that aim to protect pollinators.

Architecture and potential roles of a delta-class glutathione S-transferase in protecting honey bee from agrochemicals.

The European honey bee, Apis mellifera, serves as the principle managed pollinator species globally. In recent decades, honey bee populations have been facing serious health threats from combined biotic and abiotic stressors, including diseases, limited nutrition, and agrochemical exposure. Understanding the molecular mechanisms underlying xenobiotic adaptation of A. mellifera is critical, considering its extensive exposure to phytochemicals and agrochemicals present in the environment. In this study, we conducted a comprehensive structural and functional characterization of AmGSTD1, a delta class glutathione S-transferase (GST), to unravel its roles in agrochemical detoxification and antioxidative stress responses. We determined the 3-dimensional (3D) structure of a honey bee GST using protein crystallography for the first time, providing new insights into its molecular structure. Our investigations revealed that AmGSTD1 metabolizes model substrates, including 1-chloro-2,4-dinitrobenzene (CDNB), p-nitrophenyl acetate (PNA), phenylethyl isothiocyanate (PEITC), propyl isothiocyanate (PITC), and the oxidation byproduct 4-hydroxynonenal (HNE). Moreover, we discovered that AmGSTD1 exhibits binding affinity with the fluorophore 8-Anilinonaphthalene-1-sulfonic acid (ANS), which can be inhibited with various herbicides, fungicides, insecticides, and their metabolites. These findings highlight the potential contribution of AmGSTD1 in safeguarding honey bee health against various agrochemicals, while also mitigating oxidative stress resulting from exposure to these substances.

Systemic pesticides in a solitary bee pollen food store affect larval development and increase pupal mortality.

Solitary bees are often exposed to various pesticides applied for pest control on farmland while providing pollination services to food crops. Increasing evidence suggests that sublethal toxicity of agricultural pesticides affects solitary bees differently than the social bees used to determine regulatory thresholds, such as honey bees and bumblebees. Studies on solitary bees are challenging because of the difficulties in obtaining large numbers of eggs or young larvae for bioassays. Here we show the toxic and sublethal developmental effects of four widely used plant systemic pesticides on the Japanese orchard bee (Osmia cornifrons). Pollen food stores of this solitary bee were treated with different concentrations of three insecticides (acetamiprid, flonicamid, and sulfoxaflor) and a fungicide (dodine). Eggs were transplanted to the treated pollen and larvae were allowed to feed on the pollen stores after egg hatch. The effects of chronic ingestion of contaminated pollen were measured until adult eclosion. This year-long study revealed that chronic exposure to all tested pesticides delayed larval development and lowered larval and adult body weights. Additionally, exposure to the systemic fungicide resulted in abnormal larval defecation and increased mortality at the pupal stage, indicating potential risk to bees from fungicide exposure. These findings demonstrate potential threats to solitary bees from systemic insecticides and fungicides and will help in making policy decisions to mitigate these effects.

More than mesolectic: Characterizing the nutritional niche of Osmia cornifrons.

Characterizing the nutritional needs of wild bee species is an essential step to better understanding bee biology and providing suitable supplemental forage for at-risk species. Here, we aim to characterize the nutritional needs of a model solitary bee species, Osmia cornifrons (Radoszkowski), by using dietary protein-to-lipid ratio (P:L ratio) as a proxy for nutritional niche and niche breadth. We first identified the mean target P:L ratio (~3.02:1) and P:L collection range (0.75-6.26:1) from pollen provisions collected across a variety of sites and time points. We then investigated the P:L tolerance range of larvae by rearing bees in vitro on a variety of diets. Multifloral and single-source pollen diets with P:L ratios within the range of surveyed provisions did not always support larval development, indicating that other dietary components such as plant secondary compounds and micronutrients must also be considered in bee nutritional experiments. Finally, we used pollen metabarcoding to identify pollen from whole larval provisions to understand how much pollen bees used from plants outside of their host plant families to meet their nutritional needs, as well as pollen from individual forager bouts, to observe if bees maintained strict floral constancy or visited multiple plant genera per foraging bout. Whole larval provision surveys revealed a surprising range of host plant pollen use, ranging from ~5% to 70% of host plant pollen per provision. Samples from individual foraging trips contained pollen from multiple genera, suggesting that bees are using some form of foraging decision making. Overall, these results suggest that O. cornifrons have a wide nutritional niche breadth, but while pollen P:L ratio tolerance is broad, a tolerable P:L ratio alone is not enough to create a quality diet for O. cornifrons, and the plant species that make up these diets must also be carefully considered.

Efficacy and nontarget effects of broadcast treatments to manage spotted lanternfly (Hemiptera: Fulgoridae) nymphs.

Management to control the spotted lanternfly, Lycorma delicatula (White), would ideally achieve managers' goals while limiting impacts on nontarget organisms. In a large-scale field study with 45 plots at least 711 m2, we tested foliar applications of dinotefuran and 2 formulations of Beauveria bassiana (Balsamo) Vuillemin, each applied from the ground and separately by helicopter. Applications targeted early instar nymphs. For both application methods, a single treatment with dinotefuran significantly reduced L. delicatula numbers, as measured by catch on sticky bands (91% reduction by air and 84% reduction by ground 19 days after application) and by timed counts (89% reduction by air and 72% reduction by ground 17 days after application). None of the B. bassiana treatments significantly reduced L. delicatula numbers, even after 3 applications. Beauveria bassiana infection in field-collected nymphs ranged from 0.4% to 39.7%, with higher mortality and infection among nymphs collected from ground application plots. Beauveria bassiana conidia did not persist for long on foliage which probably contributed to low population reduction. Nontarget effects were not observed among arthropods captured in blue vane flight intercept traps, San Jose Scale pheromone sticky traps or pitfall traps, but power analysis revealed that small reductions of less than 40% may not be detected despite extensive sampling of 48,804 specimens. These results demonstrate that dinotefuran can markedly reduce local abundance of L. delicatula with little apparent effect on nontarget insects when applied shortly after hatch, and that aerial applications can match or exceed the effectiveness of applications from the ground.

Potential risk to pollinators from neonicotinoid applications to host trees for management of spotted lanternfly, Lycorma delicatula (Hemiptera: Fulgoridae).

Neonicotinoid insecticides are used to manage spotted lanternfly (Lycorma delicatula (White); hereafter SLF), a recently introduced pest in the United States. Neonicotinoids can harm nontargets, such as pollinators potentially exposed via floral resources of treated plants. We quantified neonicotinoid residues in whole flowers of two SLF host plant species, red maple (Acer rubrum L. [Sapindales: Sapindaceae]) and tree-of-heaven (Ailanthus altissima (Mill.) [Sapindales: Simaroubaceae]), treated with post-bloom imidacloprid or dinotefuran applications that differed in timing and method of application. In red maple flowers, dinotefuran residues from fall applications were significantly higher than summer applications, while imidacloprid residues from fall applications were significantly lower than summer applications. Residues did not differ between application methods or sites. In tree-of-heaven flowers, dinotefuran residues were only detected in one of 28 samples at a very low concentration. To assess acute mortality risk to bees from oral exposure to residues in these flowers, we calculated risk quotients (RQ) using mean and 95% prediction interval residue concentrations from treatments in this study and lethal concentrations obtained from acute oral bioassays for Apis mellifera (L. (Hymenoptera: Apidae)) and Osmia cornifrons (Radoszkowski (Hymenoptera: Megachilidae)), then compared these RQs to a level of concern. For A. mellifera, only one treatment group, applied at 2X maximum label rate, had an RQ that exceeded this level. However, several RQs for O. cornifrons exceeded the level of concern, suggesting potential acute risk to solitary bees. Further studies are recommended for more comprehensive risk assessments to nontargets from neonicotinoid use for SLF management.

Management of Panonychus ulmi with Various Miticides and Insecticides and Their Toxicity to Predatory Mites Conserved for Biological Mite Control in Eastern U.S. Apple Orchards.

Panonychus ulmi (Koch) (Acari: Tetranychidae), commonly known as European red mite, is a polyphagous pest of various tree and small fruit crops, including apples. A field study was conducted to evaluate different pesticide options available for the management of P. ulmi, and their impact on the population of non-target predatory mite species complex consisting of Neoseiulus fallacis, Typhlodromus pyri, and Zetzellia mali in apple orchards. Pesticides were applied using a commercial airblast sprayer at the 3-5 mite/leaf recommended economic Integrated Pest Management (IPM) threshold or prophylactically in the spring ignoring IPM practices such as monitoring, reliance on biological control and economic thresholds. Effects on the motile and egg stages of P. ulmi were evaluated as were effects on the populations of predatory mites through frequent leaf counts during the season. We also recorded the subsequent overwintering eggs of P. ulmi from each pesticide treatment. The two prophylactic treatments containing a mixture of zeta-cypermethrin + avermectin B1 + 1% horticultural oil and abamectin + 1% horticultural oil provided effective control of P. ulmi population throughout the season without reducing predatory mite populations. In contrast, eight treatments applied at the recommended economic threshold of 3-5 mites/leaf were not effective in suppressing P. ulmi populations and most reduced predatory mites. Etoxazole had significantly higher number of overwintering P. ulmi eggs compared to all other treatments.

Osmia taurus (Hymenoptera: Megachilidae): A Non-native Bee Species With Invasiveness Potential in North America.

Bees are important pollinators and are essential for the reproduction of many plants in natural and agricultural ecosystems. However, bees can have adverse ecological effects when introduced to areas outside of their native geographic ranges. Dozens of non-native bee species are currently found in North America and have raised concerns about their potential role in the decline of native bee populations. Osmia taurus Smith (Hymenoptera: Megachilidae) is a mason bee native to eastern Asia that was first reported in the United States in 2002. Since then, this species has rapidly expanded throughout the eastern part of North America. Here, we present a comprehensive review of the natural history of O. taurus, document its recent history of spread through the United States and Canada, and discuss the evidence suggesting its potential for invasiveness. In addition, we compare the biology and history of colonization of O. taurus to O. cornifrons (Radoszkowski), another non-native mason bee species now widespread in North America. We highlight gaps of knowledge and future research directions to better characterize the role of O. taurus in the decline of native Osmia spp. Panzer and the facilitation of invasive plant-pollinator mutualisms.

Six years of wild bee monitoring shows changes in biodiversity within and across years and declines in abundance.

Wild bees form diverse communities that pollinate plants in both native and agricultural ecosystems making them both ecologically and economically important. The growing evidence of bee declines has sparked increased interest in monitoring bee community and population dynamics using standardized methods. Here, we studied the dynamics of bee biodiversity within and across years by monitoring wild bees adjacent to four apple orchard locations in Southern Pennsylvania, USA. We collected bees using passive Blue Vane traps continuously from April to October for 6 years (2014-2019) amassing over 26,000 bees representing 144 species. We quantified total abundance, richness, diversity, composition, and phylogenetic structure. There were large seasonal changes in all measures of biodiversity with month explaining an average of 72% of the variation in our models. Changes over time were less dramatic with years explaining an average of 44% of the variation in biodiversity metrics. We found declines in all measures of biodiversity especially in the last 3 years, though additional years of sampling are needed to say if changes over time are part of a larger trend. Analyses of population dynamics over time for the 40 most abundant species indicate that about one third of species showed at least some evidence for declines in abundance. Bee family explained variation in species-level seasonal patterns but we found no consistent family-level patterns in declines, though bumble bees and sweat bees were groups that declined the most. Overall, our results show that season-wide standardized sampling across multiple years can reveal nuanced patterns in bee biodiversity, phenological patterns of bees, and population trends over time of many co-occurring species. These datasets could be used to quantify the relative effects that different aspects of environmental change have on bee communities and to help identify species of conservation concern.

Insect Visitors of Black Cherry (Prunus serotina) (Rosales: Rosaceae) and Factors Affecting Viable Seed Production.

Since 2000, wild black cherry (Prunus serotina Ehrh.) has produced erratic seed crops, especially in the Allegheny National Forest where poor seed production has been implicated in difficulties with black cherry regeneration in forest stands. Given that black cherry is insect pollinated and unable to produce viable seed from self-pollination, a reduction in seed crops could be due to a pollination deficit; however, its key pollinators are unknown. Identifying the pollinators and factors that influence pollinator abundance and fidelity is critical for supporting and enhancing seed production for this valuable timber species. Over a two-year period in developed, semideveloped, and forested areas in Pennsylvania, we identified the potential pollinators of black cherry and examined how their abundance, along with several other abiotic and biotic factors, influenced viable seed production. We found that andrenid (Andrenidae: Hymenoptera) bees are likely the most important pollinators. The proportion of viable seeds increased as the number of andrenids increased, and these ground nesting bees were most abundant on forest edges, highlighting this habitat's potential to support pollination services. Andrenids carried an average of 347-fold more black cherry pollen than flies and 18-fold more than halictid (Halictidae: Hymenoptera) bees. We did not find a significant relationship between the abundance of any other taxa besides andrenids and viable seed production. Black cherry flowers also provide resources for natural enemies such as the economically important parasitoid of Popillia japonica Newman (Scarabaeidae: Coleoptera), Tiphia vernalis Rohwer (Tiphiidae: Hymenoptera), which was observed feeding on black cherry nectar in this study.

CropPol: A dynamic, open and global database on crop pollination.

Seventy five percent of the world's food crops benefit from insect pollination. Hence, there has been increased interest in how global change drivers impact this critical ecosystem service. Because standardized data on crop pollination are rarely available, we are limited in our capacity to understand the variation in pollination benefits to crop yield, as well as to anticipate changes in this service, develop predictions, and inform management actions. Here, we present CropPol, a dynamic, open, and global database on crop pollination. It contains measurements recorded from 202 crop studies, covering 3,394 field observations, 2,552 yield measurements (i.e., berry mass, number of fruits, and fruit density [kg/ha], among others), and 47,752 insect records from 48 commercial crops distributed around the globe. CropPol comprises 32 of the 87 leading global crops and commodities that are pollinator dependent. Malus domestica is the most represented crop (32 studies), followed by Brassica napus (22 studies), Vaccinium corymbosum (13 studies), and Citrullus lanatus (12 studies). The most abundant pollinator guilds recorded are honey bees (34.22% counts), bumblebees (19.19%), flies other than Syrphidae and Bombyliidae (13.18%), other wild bees (13.13%), beetles (10.97%), Syrphidae (4.87%), and Bombyliidae (0.05%). Locations comprise 34 countries distributed among Europe (76 studies), North America (60), Latin America and the Caribbean (29), Asia (20), Oceania (10), and Africa (7). Sampling spans three decades and is concentrated on 2001-2005 (21 studies), 2006-2010 (40), 2011-2015 (88), and 2016-2020 (50). This is the most comprehensive open global data set on measurements of crop flower visitors, crop pollinators and pollination to date, and we encourage researchers to add more datasets to this database in the future. This data set is released for non-commercial use only. Credits should be given to this paper (i.e., proper citation), and the products generated with this database should be shared under the same license terms (CC BY-NC-SA).

Biology, Genetic Diversity, and Conservation of Wild Bees in Tree Fruit Orchards.

Different species of bees provide essential ecosystem services by pollinating various agricultural crops, including tree fruits. Many fruits and nuts depend on insect pollination, primarily by wild and managed bees. In different geographical regions where orchard crops are grown, fruit growers rely on wild bees in the farmscape and use orchard bees as alternative pollinators. Orchard crops such as apples, pears, plums, apricots, etc., are mass-flowering crops and attract many different bee species during their bloom period. Many bee species found in orchards emerge from overwintering as the fruit trees start flowering in spring, and the active duration of these bees aligns very closely with the blooming time of fruit trees. In addition, most of the bees in orchards are short-range foragers and tend to stay close to the fruit crops. However, the importance of orchard bee communities is not well understood, and many challenges in maintaining their populations remain. This comprehensive review paper summarizes the different types of bees commonly found in tree fruit orchards in the fruit-growing regions of the United States, their bio-ecology, and genetic diversity. Additionally, recommendations for the management of orchard bees, different strategies for protecting them from multiple stressors, and providing suitable on-farm nesting and floral resource habitats for propagation and conservation are discussed.

Whole-Body Acute Contact Toxicity of Formulated Insecticide Mixtures to Blue Orchard Bees (Osmia lignaria).

Blue orchard bees, [Osmia lignaria (Say) (Hymenoptera: Megachilidae)], have been developed as an important pollinator for orchard crops in North America over the last 40 years. The toxicity of several pesticides to O. lignaria and other Osmia species has been previously reported. However, the field-realistic toxicity of formulated premix insecticides comprised of multiple active ingredients (each with a different mode of action) to O. lignaria has not been assessed. Here, we use a customized spray tower in a laboratory setting to assess adult male and female whole-body direct contact exposure to four formulated pesticide mixtures: thiamethoxam + lambda-cyhalothrin (TLC), imidacloprid + beta-cyfluthrin (IBC), chlorantraniliprole + lambda-cyhalothrin (CLC) and methoxyfenozide + spinetoram (MS) by directly spraying anesthetized bees in Petri dishes. Separately, adult male and female whole-body direct contact exposure to formulated imidacloprid (I), beta-cyfluthrin (BC) and their 1:1 binary combination (IBC) was assessed using the same experimental method. Resulting mortality in each study was screened up to 96 h post-treatment to determine acute whole-body contact toxicity. In the first study, TLC and IBC resulted in statistically higher mortality at 24 and 48 h than the two other insecticide combinations tested. The CLC and MS combinations were slower acting and the highest mortality for O. lignaria exposed to these mixtures was recorded at 96 h. We did observe significant differences in toxicity between CLC and MS. In the second study, exposure to the 1:1 binary combination of IBC caused overall significantly higher mortality than exposure to I or BC alone. Both active ingredients alone, however, demonstrated equivalent levels of mortality to the 1:1 binary combination treatment at the 96 h observation reading, indicating increased speed of kill, but not necessarily increased toxicity. Significant differences in the onset of mortality following acute contact whole-body exposure to the formulated insecticide mixtures and individual active ingredients tested were consistently observed across all experiments in both studies.

Correction to 'Introduced bees (Osmia cornifrons) collect pollen from both coevolved and novel host-plant species within their family-level phylogenetic preferences'.

[This corrects the article DOI: 10.1098/rsos.200225.].

Introduced bees (Osmia cornifrons) collect pollen from both coevolved and novel host-plant species within their family-level phylogenetic preferences.

Studying the pollen preferences of introduced bees allows us to investigate how species use host-plants when establishing in new environments. Osmia cornifrons is a solitary bee introduced into North America from East Asia for pollination of Rosaceae crops such as apples and cherries. We investigated whether O. cornifrons (i) more frequently collected pollen from host-plant species they coevolved with from their geographic origin, or (ii) prefer host-plant species of specific plant taxa independent of origin. To address this question, using pollen metabarcoding, we examined the identity and relative abundance of pollen in larval provisions from nests located in different landscapes with varying abundance of East-Asian and non-Asian plant species. Our results show that O. cornifrons collected more pollen from plant species from their native range. Plants in the family Rosaceae were their most preferred pollen hosts, but they differentially collected species native to East Asia, Europe, or North America depending on the landscape. Our results suggest that while O. cornifrons frequently collect pollen of East-Asian origin, the collection of pollen from novel species within their phylogenetic familial affinities is common and can facilitate pollinator establishment. This phylogenetic preference highlights the effectiveness of O. cornifrons as crop pollinators of a variety of Rosaceae crops from different geographic origins. Our results imply that globalization of non-native plant species may ease the naturalization of their coevolved pollinators outside of their native range.

Applications of Beauveria bassiana (Hypocreales: Cordycipitaceae) to Control Populations of Spotted Lanternfly (Hemiptera: Fulgoridae), in Semi-Natural Landscapes and on Grapevines.

Spotted lanternfly, Lycorma delicatula (White), is an invasive Asian insect that was initially found in Berks County, Pennsylvania, in 2014. As of early 2020, this pest had been found in five more eastern states and it is expected to continue to expand its geographical range. Lycorma delicatula is highly polyphagous but seems to prefer tree-of-heaven, Ailanthus altissima. However, grape growers in Pennsylvania have reported significant damage and loss of vines caused by L. delicatula adults. In fall 2018, two fungal entomopathogens (Beauveria bassiana and Batkoa major) drove localized collapses in L. delicatula populations in Berks County, Pennsylvania. In 2019, we tested applications of a commercialized mycoinsecticide based on B. bassiana strain GHA on L. delicatula populations in a public park in southeastern Pennsylvania. A single application of B. bassiana reduced fourth instar nymphs by 48% after 14 d. Applications of B. bassiana to L. delicatula adults in the same park resulted in 43% mortality after 14 d. Beauveria bassiana spores remained viable on foliage for 5-7 d after spraying. We also conducted semi-field bioassays with B. bassiana GHA (formulated as BoteGHA and Aprehend) and another mycoinsecticide containing Isaria fumosorosea Apopka Strain 97 against L. delicatula adults feeding on potted grapes. All the mycoinsecticides killed ≥90% of adults after 9 d using direct applications. Aprehend killed 99% of adults after 9 d with exposure to residues on sprayed grapes. These data show that fungal entomopathogens can help to suppress populations of L. delicatula in agroecosystems and natural areas.

A new ingestion bioassay protocol for assessing pesticide toxicity to the adult Japanese orchard bee (Osmia cornifrons).

Adopting an Integrated Pest and Pollinator Management strategy requires an evaluation of pesticide risk for pollinator species. For non-Apid species, however, the standardized ingestion assays are difficult to implement. This hinders the consideration of non-Apid species in farm management strategies and government regulatory processes. We describe a new method for a mason bee, Osmia cornifrons, which is an important pollinator of apples and other fruit crops. Our approach overcomes high control mortality seen in other bioassay protocols and expands testing to include males as well as females. The new pesticide toxicity assessment protocol showed that (1) a group feeding method is optimum even though there is no trophallaxis, (2) males had better tolerance to pesticides although they are smaller, and (3) pesticides can cause additional mortality after the standard 48 h assessment time specified by European Food Safety Authority and U.S. Environmental Protection Agency.

A Method for a Long-Term Marking of Spotted Lanternfly (Hemiptera: Fulgoridae) Using a Stable Isotope of Nitrogen.

Developing a lifelong marking method for Lycorma delicatula (White, 1845) is crucial to investigate ecological processes. Here we validate a marking method using stable isotope enrichment (15N) of host plants to track spotted lanternfly (SLF), an invasive species causing economic damage on grapes, hardwood forest and landscape tree species. To validate this method, we first determined the isotope dosage to be sprayed on the host plants and subsequently detected in SLF. Second, we examined whether 15N mark remains detectable from the nymphal to adult stage. We demonstrated that two stable isotope dosages applied to the host plants were assimilated by the insect and equally detectable in the exoskeleton, wings, and mature eggs ready to be oviposited. This safe and reliable method can be used to examine fundamental processes of the biology and ecology of SLF that range from dietary resources and resource allocation to food-web structure and dispersal patterns.