Agri-environment scheme nectar chemistry can suppress the social epidemiology of parasites in an important pollinator.

Reinventing the plant

To accurately estimate exposure of bees to pesticides, analytical methods are needed to enable quantification of nanogram/gram (ng/g) levels of contaminants in small samples of pollen or the individual insects. A modified QuEChERS extraction method coupled with ultra-high-performance liquid chromatography tandem mass spectrometry (UHPLC-MS/MS) analysis was tested to quantify residues of 19 commonly used neonicotinoids and fungicides and the synergist, piperonyl butoxide, in 100mg samples of pollen and in samples of individual bumblebees (Bombus terrestris). Final recoveries ranged from 71 to 102% for most compounds with a repeatability of below 20% for both pollen and bumblebee extracts spiked at 5 and 40ng/g. The method enables the detection of all compounds at sub-ng/g levels in both matrices and the method detection limits (MDL) ranged from 0.01 to 0.84ng/g in pollen and 0.01 to 0.96ng/g in individual bumblebees. Using this method, mixtures of neonicotinoids (thiamethoxam, clothianidin, imidacloprid and thiacloprid) and fungicides (carbendazim, spiroxamine, boscalid, tebuconazole, prochloraz, metconazole, fluoxastrobin, pyraclostrobin and trifloxystrobin) were detected in pollens of field bean, strawberry and raspberry at concentrations ranging from <MDL to 67ng/g for neonicotinoids and from <MDL to 14ng/g for fungicides. In bumblebees, the insecticides thiamethoxam and thiacloprid were present at concentrations >MDL, and in some bees, the fungicides carbendazim, boscalid, tebuconazole, flusilazole and metconazole were present at concentrations between 0.80 to 30ng/g. This new method allows the analysis of mixtures of neonicotinoids and fungicides at trace levels in small quantities of pollen and individual bumblebees and thus will facilitate exposure assessment studies.

The objective of this study was to quantify whether the presence of three different neonicotinoid insecticides (imidacloprid, thiamethoxam or clothianidin) in sucrose solution results in antifeedant effects in individual worker bumblebees (Bombus terrestris), and, if so, whether this effect is reversible if bees are subsequently offered untreated feed. Bees exposed to imidacloprid displayed a significant dose-dependent reduction in consumption at 10 and 100 µg L(-1), which was reversed when untreated feed was offered. No consistent avoidance/antifeedant response to nectar substitute with thiamethoxam was detected at the more field-realistic dose rates of 1 and 10 µg L(-1), and exposure to the very high 100 µg L(-1) dose rate was followed by 100% mortality of experimental insects. No reduction in food intake was recorded at 1 µg clothianidin L(-1), reduced consumption was noted at 10 µg clothianidin L(-1) and 100% mortality occurred when bees were exposed to rates of 100 µg clothianidin L(-1). This study provides evidence of a direct antifeedant effect of imidacloprid and clothianidin in individual bumblebees but highlights that this may be a compound-specific effect.

An important function of agri-environmental schemes (AES) is to change management of pastures to better conserve biodiversity. However, the effects of most AES on biodiversity are poorly understood, especially when it comes to effects of AES management over time. The main aim of this study is to investigate if the species richness and abundance of grassland specialists of vascular plants and two important insect pollinator groups (bumblebees and butterflies) differ over time (5 years) in pastures with AES management (two value levels; general values and special values) and pastures without AES management. We also investigate if local vegetation characteristics and landscape composition relate to species richness in semi-natural grasslands. Using data from more than 400 sites we found that species richness of vascular plants (grassland specialists) was higher in pastures with AES management (for special and general values) compared to those without AES, which implies that these schemes do have value of the conservation of plant diversity. However, species richness and abundance of butterflies (grassland specialists) and bumblebees (all species) did not differ significantly among the three AES categories. We found no evidence that the type of AES management caused any changes in species richness of plants, butterflies or bumblebees during the 5 year period of our investigation. It appears that AES management that encourages uniform and minimum levels of grazing can have both positive and negative effects on biodiversity. For example, pollinators may benefit from a lower grazing intensity that could increase flower richness and heterogeneity in vegetation height. However, low grazing intensity may lead to increased cover of trees and shrubs, which can have negative effects for both insect pollinators and vascular plants. The effects of landscape composition were weak and only species richness of bumble bees were associated with landscape composition. Designing management regimes to maintain suitably heterogeneous vegetation layer, and continued long-term monitoring of biodiversity will be critical for safeguarding culturally and functionally important semi-natural grasslands.

One key target of agri-environmental schemes (AES) is maintaining biodiversity. However, if and how insect biomass and diversity benefit from grassland AES is not well known. Using Malaise trap samples from 72 grassland sites in four grassland dominated regions in Southern Germany, we investigated the effect of 1) local grassland AES (three levels: control, farm-wide, site-specific), and 2) an index of land use extensification (LEI, calculated for each site) on insect biomass and species richness of all insects, of five insect orders including all species (Coleoptera, Diptera, Hemiptera, Hymenoptera, Lepidoptera), and two pollinator groups (Syrphidae, Apiformes). Moreover, the effect of grassland AES in the surrounding landscape was quantified. Insect samples were subjected to metabarcoding to create DNA barcode sequences which were then compared with reference libraries (BOLD; NCBI) and additionally classified using the Ribosomal Database Project (RDP) classifier. Resulting taxonomies were combined into a consensus taxonomy table and the potential minimum number of species per sample was estimated. Statistical analyses were carried out using Generalized linear mixed models and Bayesian inference. Our results indicated that species richness of many insect groups benefited from AES. As predicted by the regression model, presence of site-specific grassland AES increased the species richness of all insects (∼8%), Hymenoptera (∼27%), Lepidoptera (∼35%) and Syrphidae (∼26%). While Apiformes benefited most, we found an average increase of species richness in the presence of farm-wide AES by ∼82% and even more with site-specific AES ∼120%. We found that an increasing LEI, and thus more extensive management, positively affected insect diversity in grassland habitats. No effect of local AES and LEI on insect biomass was found. Moreover, the amount of grassland AES in the landscape was less important for insect diversity. We conclude that many grassland insects (especially pollinators) benefit from extensive management. However, we showed that the studied AES variants comprise variable management intensities hampering general AES recommendations for promoting insects. AES programs should, therefore, specifically reward extensification measures that were shown to benefit insect diversity. The integration of the land use extensification index (LEI) as a tool in the AES framework seems a promising approach to preserve and promote of insect diversity.

The northern section of the Caspian Sea and lower reaches of the Zhaiyk (Ural) River is an important fishery for Kazakhstan. In the present study, a total of 1597 individuals of ten fish species were analysed. The fish were caught over three years, from 2018 to 2020. For each species studied - Abramis brama, Alosa saposchnikowii, Atherina boyeri caspia, Carassius gibelio, Chelon auratus, Cyprinus carpio, Leuciscus aspius, Rutilus caspius, Sander marinus and Sander volgensis - between 100 and 200 individuals were examined. A series of generalized linear models (GLMs) were used to examine the association between individual parasite intensity of infection and the Fulton index, age, year the fish was captured, where the fish was captured (northern Caspian or Zhaiyk River) and sex. For each GLM, the best-fitting probability distribution was used -either Poisson, zero-inflated Poisson, negative binomial or zero-inflated negative binomial. For some fish/parasite species, an increased Fulton index was associated with higher intensities of parasite infection, whilst, for others, the Fulton index decreased with the intensity of parasite infection. This was also true of age-related intensity of infection, with some parasites having an increased intensity of infection with age whilst others had a decreased intensity of infection with age. There was also some evidence of variation in intensity of parasite infection between different years when the fish were caught. For some species of fish that are endemic to both the fresh waters of the Zhaiyk River and the low-saline waters of the northern Caspian, there were variations in intensity of parasite infection between the two environments. The best-fitting probability distribution also gave some information about the dynamics of infection. No fish species had a Poisson distribution of parasites, which is consistent with an entirely random infection process, with all fish being potentially exposed. For some parasites, the distribution was a zero-inflated Poisson, which is consistent with either the fish being exposed to parasite infection or not; and, if exposed, infection was a random process. Other parasites had a negative binomial distribution, consistent with the entire fish population being exposed, but the infection process was clumped or there were variations in the susceptibility of infection between fish. Finally, some of the parasites had a zero-inflated negative binomial distribution, which can be interpreted as part of the fish population not being exposed and the remainder of the population being exposed to a clumped or aggregated infection process and/or a variation in individual susceptibility to infection.

Body mass and mate choice in bumblebees (Bombus terrestris) under climate heating.

Simple SummaryHoneybees (Apis mellifera) and bumblebees (Bombus terrestris) are often used to pollinate melon flowers in facilities. The researchers identified the volatiles in male and female flowers of a common melon species (Cucumis melo) grown in facilities and measured the EAG and behavioral responses of honeybees and bumblebees when exposed to isolated volatiles from the melon flowers. These results provide basic data for the rational utilization of bees.As important pollinators, honeybees and bumblebees present a pollination behavior that is influenced by flower volatiles through the olfactory system. In this study, volatile compounds from melon flowers were isolated and identified by headspace solid-phase microextraction (HS-SPME) and gas chromatography–mass spectrometry (GC-MS), and their effects on Apis mellifera and Bombus terrestris were investigated by electroantennogram (EAG) and behavior tests (Y-tube olfactometer). The results showed that 77 volatile compounds were detected in melon flowers, among which the relative content of aldehydes was the highest (61.34%; 82.09%). A. mellifera showed a strong EAG response to e-2-hexenal, e-2-octenal, and 1-nonanal. B. terrestris showed a strong EAG response to e-2-hexenal, e-2-octenal, 2,5-dimethyl-benzaldehyde, benzaldehyde and benzenepropanal. In behavior tests, the volatiles with the highest attractive rate to A. mellifera were e-2-hexenal (200 μg/μL, 33.33%) and e-2-octenal (300 μg/μL, 33.33%), and those to B. terrestris were e-2-hexenal (10 μg/μL, 53.33%) and 2,5-dimethyl-benzaldehyde (100 μg/μL, 43.33%). E-2-hexenal and e-2-octenal were more attractive to A. mellifera than B. terrestris, respectively (10 μg/μL, 10 μg/μL, 200 μg/μL). In conclusion, the volatiles of melon flowers in facilities have certain effects on the electrophysiology and behavior of bees, which is expected to provide theoretical and technical support for the pollination of A. mellifera and B. terrestris in facilities.

Individual bumblebees were trained to choose between rewarded target flowers and non-rewarded distractor flowers in a controlled illumination laboratory. Bees learnt to discriminate similar colours, but with smaller colour distances the frequency of errors increased. This indicates that pollen transfer might occur between flowers with similar colours, even if these colours are distinguishable. The effect of similar colours on reducing foraging accuracy of bees is evident for colour distances high above discrimination threshold, which explains previous field observations showing that bees do not exhibit complete flower constancy unless flower colour between species is distinct. Bees tested in spectrally different illumination conditions experienced a significant decrease in their ability to discriminate between similar colours. The extent to which this happens differs in different areas of colour space, which is consistent with a von Kries-type model of colour constancy. We find that it would be beneficial for plant species to have highly distinctive colour signals to overcome limitations on the bees performance in reliably judging differences between similar colours. An exception to this finding was flowers that varied in shape, in which case bees used this cue to compensate for inaccuracies of colour vision.

Widespread use of neonicotinoid insecticides, such as imidacloprid, is often associated with diminishing populations of bees; this loss of pollinators presents a concern for food security and may cause unpredictable changes in ecological networks. However, little is known about the potential behavioural mechanisms behind the neonicotinoid-associated pollinator decline. We quantified the effects of low-dose (1 ppb) imidacloprid exposure on the foraging behaviour of bumblebees (Bombus terrestris). Individual bumblebees were released into a flight arena containing three patches of robotic flowers whose colour (yellow, orange, blue) indicated whether the flower delivered a reward (sugar solution). Exposure to imidacloprid had no significant effect on measures of bumblebee physical performance (such as flight speed) or learning (identifying rewarding flowers). However, pesticide-treated bumblebees had reduced foraging motivation compared with the control bumblebees, as they visited fewer robotic flowers, were slower to start foraging and did not visit all three flower colours as often. Neonicotinoid concentrations of 1 ppb, often reported in plant nectar near agricultural lands, can thus affect the foraging behaviour of bumblebees. Even without a notable impact on flight performance and learning, a reduction in foraging motivation could explain the poor performance of colonies of bumblebees exposed to neonicotinoids.

Israeli acute paralysis virus (IAPV), a single-stranded RNA virus, has a worldwide distribution and affects honeybees as well as other important pollinators. IAPV infection in honeybees has been successfully repressed by exploiting the RNA interference (RNAi) pathway of the insect’s innate immune response with virus-specific double stranded RNA (dsRNA). Here we investigated the effect of IAPV infection in the bumblebee Bombus terrestris and its tissue tropism. B. terrestris is a common pollinator of wild flowers in Europe and is used for biological pollination in agriculture. Infection experiments demonstrated a similar pathology and tissue tropism in bumblebees as reported for honeybees. The effect of oral administration of virus-specific dsRNA was examined and resulted in an effective silencing of the virus, irrespective of the length. Interestingly, we observed that non-specific dsRNA was also efficient against IAPV. However further study is needed to clarify the precise mechanism behind this effect. Finally we believe that our data are indicative of the possibility to use dsRNA for a broad range viral protection in bumblebees.

Infection with the trypanosome Crithidia bombi and expression of immune-related genes in the bumblebee Bombus terrestris

SummaryNeonicotinoids have been implicated in the large declines observed in insects such as bumblebees, an important group of pollinators. Neonicotinoids are agonists of nicotinic acetylcholine receptors that are found throughout the insect central nervous system and are the main mediators of synaptic neurotransmission. These receptors are important for the function of the insect central clock and circadian rhythms. The clock allows pollinators to coincide their activity with the availability of floral resources and favorable flight temperatures, as well as impact learning, navigation, and communication. Here we show that exposure to the field-relevant concentration of 10 μg/L imidacloprid caused a reduction in bumblebee foraging activity, locomotion, and foraging rhythmicity. Foragers showed an increase in daytime sleep and an increase in the proportion of activity occurring at night. This could reduce foraging and pollination opportunities, reducing the ability of the colony to grow and reproduce, endangering bee populations and crop yields.

Simple SummaryTomatoes are a popular crop, and bumble bees and honey bees are its main pollinators. Floral scent usually plays an important role in mediating the foraging behavior of bees, and tomato flowers release special scents. Although it has been found that foraging experience in the tomato greenhouses helped bumble bees develop a strong preference for the scent, honey bees with foraging experience continued to show aversion to tomato floral scent. However, it is currently unknown as to how a single tomato volatile compound regulates the foraging behavior of bees. In the current study, we investigated the foraging behaviors of the widely used pollinator honey bee Apis mellifera and bumble bee Bombus terrestris on tomato flower volatile compounds in order to evaluate whether honey bees and bumble bees show different EAG responses to volatile compounds and how they might influence bee choice behavior. We found that honey bees had a weaker EAG response to the tested compounds compared with bumble bees and that they showed avoidance behavior to these compounds. We conclude that some compounds in tomato floral scents caused the low bias of honey bees to tomato flowers, which may be one driver as to why honey bees dislike tomato, which could be adjusted in order to improve the pollination service efficiency of bees for commercial crops.Bumble bees and honey bees are of vital importance for tomato pollination, although honey bees are less attracted to tomato flowers than bumble bees. Little is known about how tomato flower volatile compounds influence the foraging behaviors of honey bees and bumble bees. In this study, compounds of tomato flower volatiles were detected by gas chromatography–mass spectrometry. Electroantennography (EAG) and a dynamic two-choice olfactometer were used, respectively, to compare the differences of antennal and behavioral responses between Apis mellifera and Bombus terrestris towards selected volatile compounds. A total of 46 compounds were detected from the tomato flower volatiles. Of the 16 compounds tested, A. mellifera showed strong antennal responses to 3 compounds (1-nonanal, (+)-dihydrocarvone, and toluene) when compared with a mineral oil control, and B. terrestris showed 7 pronounced EAG responses (1,3-xylene, (+)-dihydrocarvone, toluene, piperitone, eucarvone, 1-nonanal, and β-ocimene). Additionally, 1-nonanal and (+)-dihydrocarvone elicited significant avoidance behavior of A. mellifera, but not of B. terrestris. In conclusion, bumble bees are more sensitive to the compounds of tomato flower volatiles compared to honey bees, and honey bees showed aversion to some compounds of tomato flower volatiles. The findings indicated that compounds of flower volatiles significantly influenced bee foraging preference for tomato.

Summary The primary aim of many agri‐environment schemes (AES) is to enhance biodiversity; however, the results of AES designed for this purpose have, to date, been largely underwhelming. One reason for this may be because AES tend to be administered at the farm scale. We argue that collaborative AES (cAES) – single environmental management agreements for multiple farm units – can benefit biodiversity. We then discuss how a shift to this type of scheme may impact upon ecosystem services (ES). Evidence gathered from studies carried out across European farmland demonstrated that more than a third (18 of 52; 35%) of the bird, mammal, reptile, amphibian and bumblebee species important in English farmland operate at scales larger than the typical English farm (146 ha) in the breeding season. While this study relates ranging behaviour specifically to England and the English AES system, the estimates of ranging behaviour presented in this study could also be used (with caution) to compare with farm sizes elsewhere in Europe, making the results of wider geographical relevance. Data were obtained from face‐to‐face interviews with 32 farmers with differing AES backgrounds [current participants at either low or high level (entry‐level Stewardship or higher‐level Stewardship/countryside Stewardship scheme), or current non‐participants]. Eighty‐one per cent of interviewees were found to be willing, in principle, to participate in a collaborative AES (cAES) programme. However, they viewed less extensive options (e.g. management of existing hedgerows) more favourably than extensive or cooperatively demanding land management options (e.g. large‐scale habitat creation). Data from an online consultation of a further 122 farmers supported the principal finding, with 75% of respondents willing, in theory, to participate in collaborative schemes. Synthesis and applications. Well‐designed landscape‐scale schemes are likely to be more beneficial than farm‐scale schemes for a small but significant number of key farmland species and ES, such as bats, mammals and some important pollinators, while unlikely to harm species operating at smaller scales. These schemes can be expected to attract widespread participation from landowners. Thus, policymakers may be heartened that collaborative AES are a potential multifaceted solution to environmental management on farmland.