Flight performance of pollen starved honey bees and incomplete compensation through ingestion after early life pollen deprivation.

We investigated the effect of adult honey bee pollen nutrition on the flight performance of honey bees. Therefore, caged bees were allowed to perform 30 min of defecation/training flights every second day before flight performance of pollen-fed bees and pollen-deprived bees older than 16 days were compared in a flight mill. We first fed 10 µL of 1 M glucose solution to bees, and after they metabolized this during flight, they were fed 10 µL of 2 M glucose solution for a second flight test. Pollen-deprived bees flew longer and further than pollen-fed bees in both flights. Pollen-fed bees flew faster in the early period at the beginning of flights, whereas pollen-deprived bees were faster in the final phases. Pollen-fed bees were able to raise their maximum flight speed in 2 M glucose solution flights, whereas pollen-constraint bees were not. The two groups did not differ in abdomen fresh weight, but the fresh weight of the head and thorax and dry weight of the head, thorax and abdomen were higher in pollen-fed bees. In a second experiment, we constrained pollen consumption of caged bees during the first 7 days and compared daily consumption of bees from day 8-16 to consumption of bees unrestricted in pollen. We found that pollen-deprived bees perceive the pollen shortage and try to compensate for their needs by consuming significantly more pollen at the later phase of their life than pollen-fed bees of the same age. Still, bees constrained from pollen in the first 7 days did only reach 51.1% of the lifetime consumption of unconstrained bees. This shows that bees can sense the need for essential nutrients from pollen, but their physiological apparatus does not allow them to fully compensate for their early life constraint. Pollen deprivation only in the first 7 days of worker life likewise significantly reduced fresh and dry weights of the body sections (head, thorax, and abdomen) and survival. This underlines the importance of protein consumption in a short critical period early in adult bees' lives for their development and their performance later in life.

CSI Pollen: Diversity of Honey Bee Collected Pollen Studied by Citizen Scientists.

A diverse supply of pollen is an important factor for honey bee health, but information about the pollen diversity available to colonies at the landscape scale is largely missing. In this COLOSS study, beekeeper citizen scientists sampled and analyzed the diversity of pollen collected by honey bee colonies. As a simple measure of diversity, beekeepers determined the number of colors found in pollen samples that were collected in a coordinated and standardized way. Altogether, 750 beekeepers from 28 different regions from 24 countries participated in the two-year study and collected and analyzed almost 18,000 pollen samples. Pollen samples contained approximately six different colors in total throughout the sampling period, of which four colors were abundant. We ran generalized linear mixed models to test for possible effects of diverse factors such as collection, i.e., whether a minimum amount of pollen was collected or not, and habitat type on the number of colors found in pollen samples. To identify habitat effects on pollen diversity, beekeepers' descriptions of the surrounding landscape and CORINE land cover classes were investigated in two different models, which both showed that both the total number and the rare number of colors in pollen samples were positively affected by 'urban' habitats or 'artificial surfaces', respectively. This citizen science study underlines the importance of the habitat for pollen diversity for bees and suggests higher diversity in urban areas.

Prisoners receive food fit for a queen: honeybees feed small hive beetles protein-rich glandular secretions through trophallaxis.

The honeybee nest parasite Aethina tumida (small hive beetle) uses behavioural mimicry to induce trophallactic feeding from its honeybee hosts. Small hive beetles are able to induce honeybee workers to share the carbohydrate-rich contents of their crops, but it is not clear whether the beetles are able to induce to workers to feed them the protein-rich hypopharyngeal glandular secretions fed to the queen, larvae and other nest mates. Protein is a limiting macronutrient in an insect's diet, essential for survival, growth and fecundity. Honeybees obtain protein from pollen, which is consumed and digested by nurse bees. They then distribute the protein to the rest of the colony in the form of hypopharyngeal gland secretions. Using 14C-phenylalanine as a qualitative marker for protein transfer, we show that small hive beetles successfully induce worker bees to feed them the protein-rich secretions of their hypopharyngeal glands during trophallaxis, and that females are more successful than males in inducing the transfer of these protein-rich secretions. Furthermore, behavioural observations demonstrated that female beetles do not preferentially interact with a specific age cohort of bees when soliciting food, but males tend to be more discriminant and avoid the more aggressive and active older bees.

A citizen science supported study on seasonal diversity and monoflorality of pollen collected by honey bees in Austria.

Austrian beekeepers participated in the "C.S.I. Pollen" study as citizen scientists and collected pollen from honey bee colonies in hive mounted traps every three weeks from April to September in 2014 and 2015 to uncover the seasonal availability of pollen sources for bees. 1622 pollen samples were collected and analysed using palynological light microscopy to the lowest taxonomic level possible. For 2014 and 2015 combined, 239 pollen types from more than 85 families were detected. 'Various unknown' species, Taraxacum-form and Plantago spp. were the pollen types collected by the majority of colonies (occurrence), whereas the most pollen grains collected were from Trifolium repens-form, Plantago spp. and Salix spp. (abundance). In spring, trees were found to be the most abundant pollen source, whereas in summer herbs dominated. On average, a colony collected pollen from 16.8 ± 4.7 (2014) and 15.0 ± 4.4 (2015) pollen types per sampling. Those numbers, however, vary between sampling dates and indicate a seasonal pattern. This is also supported by Simpson's diversity index, which was on median 0.668. In both years, 50.0% of analysed pollen samples were partially (>50%) and 4.2% were highly monofloral (i.e. containing >90% of one pollen type). Prevalence of monofloral pollen samples peaked at the beginning and the end of the season, when pollen diversity was the lowest.

Health status of honey bee colonies (Apis mellifera) and disease-related risk factors for colony losses in Austria.

Austrian beekeepers frequently suffered severe colony losses during the last decade similar to trends all over Europe. This first surveillance study aimed to describe the health status of Austrian bee colonies and to analyze the reasons for losses for both the summer and winter season in Austria. In this study 189 apiaries all over Austria were selected using a stratified random sampling approach and inspected three times between July 2015 and spring 2016 by trained bee inspectors. The inspectors made interviews with the beekeepers about their beekeeping practice and the history of the involved colonies. They inspected a total of 1596 colonies for symptoms of nine bee pests and diseases (four of them notifiable diseases) and took bee samples for varroa mite infestation analysis. The most frequently detected diseases were three brood diseases: Varroosis, Chalkbrood and Sacbrood. The notifiable bee pests Aethina tumida and Tropilaelaps spp. were not detected. During the study period 10.8% of the 1596 observed colonies died. Winter proved to be the most critical season, in which 75% of the reported colony losses happened. Risks for suffering summer losses increased significantly, when colonies were weak in July, had queen problems or a high varroa mite infestation level on bees in July. Risks for suffering winter losses increased significantly, when the colonies had a high varroa mite infestation level on bees in September, were weak in September, had a queen older than one year or the beekeeper had few years of beekeeping experience. However, the effect of a high varroa mite infestation level in September had by far the greatest potential to raise the winter losses compared to the other significant factors.

Investigating the role of landscape composition on honey bee colony winter mortality: A long-term analysis.

The health of honey bee colonies is, amongst others, affected by the amount, quality and diversity of available melliferous plants. Since landscape is highly diverse throughout Austria regarding the availability of nutritional resources, we used data from annual surveys on honey bee colony losses ranging over six years to analyse a possible relationship with land use. The data set comprises reports from a total of 6,655 beekeepers and 129,428 wintered honey bee colonies. Regions surrounding the beekeeping operations were assigned to one of six clusters according to their composition of land use categories by use of a hierarchical cluster analysis, allowing a rough distinction between urban regions, regions predominated by semi-natural areas and pastures, and mainly agricultural environments. We ran a Generalised Linear Mixed Model and found winter colony mortality significantly affected by operation size, year, and cluster membership, but also by the interaction of year and cluster membership. Honey bee colonies in regions composed predominantly of semi-natural areas, coniferous forests and pastures had the lowest loss probability in four out of six years, and loss probabilities within these regions were significantly lower in five out of six years compared to those within regions composed predominantly of artificial surfaces, broad-leaved and coniferous forest.

Does the Pollen Diet Influence the Production and Expression of Antimicrobial Peptides in Individual Honey Bees?

We investigated the importance of protein nutrition for honey bee immunity. Different protein diets (monofloral pollen of Helianthus spp., Sinapis spp., Asparagus spp., Castanea spp., a mixture of the four different pollen and the pollen substitute FeedbeeTM) were fed to honey bees in cages ad libitum. After 18 days of feeding, apidaecin 1 isoforms concentration in the thorax were measured using nanoflow liquid chromatography coupled with mass spectrometry. Expression levels of genes, coding for apidaecins and abaecin in the abdomen were determined using quantitative PCR. The results indicate that protein-containing nutrition in adult worker honey bees can trigger certain metabolic responses. Bees without dietary protein showed lower apidaecin 1 isoforms concentrations. The significantly lowest concentration of apidaecin 1 isoforms was found in the group that was fed no pollen diet when compared to Asparagus, Castanea, Helianthus, and Sinapis pollen or the pollen supplement FeedBeeTM. Expression levels of the respective genes were also affected by the protein diets and different expression levels of these two antimicrobial peptides were found. Positive correlation between concentration and gene expression of apidaecins was found. The significance of feeding bees with different protein diets, as well as the importance of pollen nutrition for honey bee immunity is demonstrated.

Protein nutrition governs within-host race of honey bee pathogens.

Multiple infections are common in honey bees, Apis mellifera, but the possible role of nutrition in this regard is poorly understood. Microsporidian infections, which are promoted by protein-fed, can negatively correlate with virus infections, but the role of protein nutrition for the microsporidian-virus interface is unknown. Here, we challenged naturally deformed wing virus - B (DWV-B) infected adult honey bee workers fed with or without pollen ( = protein) in hoarding cages, with the microsporidian Nosema ceranae. Bee mortality was recorded for 14 days and N. ceranae spore loads and DWV-B titers were quantified. Amongst the groups inoculated with N. ceranae, more spores were counted in protein-fed bees. However, N. ceranae infected bees without protein-diet had reduced longevity compared to all other groups. N. ceranae infection had no effect on protein-fed bee's longevity, whereas bees supplied only with sugar-water showed reduced survival. Our data also support that protein-feeding can have a significant negative impact on virus infections in insects. The negative correlation between N. ceranae spore loads and DWV-B titers was stronger expressed in protein-fed hosts. Proteins not only enhance survival of infected hosts, but also significantly shape the microsporidian-virus interface, probably due to increased spore production and enhanced host immunity.

Cuticular hydrocarbon cues of immune-challenged workers elicit immune activation in honeybee queens.

Recently, evidence has shown that variations in the cuticular hydrocarbons (CHCs) profile allow healthy honeybees to identify diseased nestmates, eliciting agonistic responses in the former. Here, we determined whether these 'immunologic cues' emitted by diseased nestmates were only detected by workers, who consequently took hygienic measures and excluded these individuals from the colony, or whether queens were also able to detect these cues and respond accordingly. Healthy honeybee queens were exposed to (i) healthy, (ii) Ringer-injected and (iii) lipopolysaccharide (LPS)-injected nestmates by allowing direct body contact. Quantitative differences in the CHC profiles of these three groups were measured using GC-MS. The transcript levels of the products of four genes that encode for antimicrobial peptides (AMPs), which are part of the queen's immune response, were measured in bees exposed to direct contact using qPCR. A significant increase in the transcript levels of these AMP genes over baseline levels in queens was observed when body contact was allowed between the queens and the Ringer- and LPS-injected nestmates. These results provide the first evidence that the detection of CHCs contributes to the initiation of an immune response in insects. In an additional experiment, CHCs were extracted from diseased workers and directly presented to queens, which also evoked a similar immune response. A potential mechanism that relied on volatile compounds could be ruled out by conducting a distance experiment. The study helps to expand our knowledge of chemical communication in insects and sheds light on a likely new mechanism of social immunity.

Food consumption and food exchange of caged honey bees using a radioactive labelled sugar solution.

We measured the distribution of sugar solution within groups of caged honey bees (Apis mellifera) under standard in vitro laboratory conditions using 14C polyethylene glycol as a radioactive marker to analyze ingestion by individual bees after group feeding. We studied the impact of different experimental setups by varying the number of bees, age of bees, origin of bees, duration of experiment, the amount of available diet, and the influence of the neurotoxic pesticide imidacloprid in the diet on the feeding and food sharing behavior (trophallaxis). Sugar solution was non-uniformly distributed in bees in 36 out of 135 cages. As a measure of the extent to which the sugar diet was equally distributed between caged bees, we calculated the (inner 80%) intake ratio by dividing the intake of the 90th percentile bee by the intake of the 10th percentile bee. This intake ratio ranged from 1.3 to 94.8 in 133 individual cages, further supporting a non-uniform distribution of food among caged bees. We can expect a cage with 10 or 30 bees containing one bee that ingests, on average, the 8.8-fold of the bee in the same cage ingesting the smallest quantity of food. Inner 80% intake ratios were lower in experiments with a permanent or chronic offering of labelled sugar solution compared to temporary or acute feedings. After pooling the data of replicates to achieve a higher statistical power we compared different experimental setups. We found that uniform food distribution is best approached with 10 newly emerged bees per cage, which originate from a brood comb from a single colony. We also investigated the trophallaxis between caged honey bees which originally consumed the diet and newly added bees. Color marked bees were starved and added to the cages in a ratio of 10:5 or 20:20 after the initial set of bees consumed all the labelled sugar solution. The distribution of the labelled sugar solution by trophallaxis within 48 hours to added bees was 25% (10:5) or 45% (20:20) of the initial sugar solution. Imidacloprid at its median lethal dose (LD50) in the sugar solution reduced this post-feeding food transmission to 27% (20:20). Our results show that differences in food intake exist within caged bees that may lead to differential exposure that can influence the interpretation of toxicity tests.

Sublethal pesticide doses negatively affect survival and the cellular responses in American foulbrood-infected honeybee larvae.

Disclosing interactions between pesticides and bee infections is of most interest to understand challenges that pollinators are facing and to which extent bee health is compromised. Here, we address the individual and combined effect that three different pesticides (dimethoate, clothianidin and fluvalinate) and an American foulbrood (AFB) infection have on mortality and the cellular immune response of honeybee larvae. We demonstrate for the first time a synergistic interaction when larvae are exposed to sublethal doses of dimethoate or clothianidin in combination with Paenibacillus larvae, the causative agent of AFB. A significantly higher mortality than the expected sum of the effects of each individual stressor was observed in co-exposed larvae, which was in parallel with a drastic reduction of the total and differential hemocyte counts. Our results underline that characterizing the cellular response of larvae to individual and combined stressors allows unmasking previously undetected sublethal effects of pesticides in colony health.

Modelling seasonal effects of temperature and precipitation on honey bee winter mortality in a temperate climate.

Insect pollinators are essential to global food production. For this reason, it is alarming that honey bee (Apis mellifera) populations across the world have recently seen increased rates of mortality. These changes in colony mortality are often ascribed to one or more factors including parasites, diseases, pesticides, nutrition, habitat dynamics, weather and/or climate. However, the effect of climate on colony mortality has never been demonstrated. Therefore, in this study, we focus on longer-term weather conditions and/or climate's influence on honey bee winter mortality rates across Austria. Statistical correlations between monthly climate variables and winter mortality rates were investigated. Our results indicate that warmer and drier weather conditions in the preceding year were accompanied by increased winter mortality. We subsequently built a statistical model to predict colony mortality using temperature and precipitation data as predictors. Our model reduces the mean absolute error between predicted and observed colony mortalities by 9% and is statistically significant at the 99.9% confidence level. This is the first study to show clear evidence of a link between climate variability and honey bee winter mortality.

How a life-like system emerges from a simple particle motion law.

Self-structuring patterns can be observed all over the universe, from galaxies to molecules to living matter, yet their emergence is waiting for full understanding. We discovered a simple motion law for moving and interacting self-propelled particles leading to a self-structuring, self-reproducing and self-sustaining life-like system. The patterns emerging within this system resemble patterns found in living organisms. The emergent cells we found show a distinct life cycle and even create their own ecosystem from scratch. These structures grow and reproduce on their own, show self-driven behavior and interact with each other. Here we analyze the macroscopic properties of the emerging ecology, as well as the microscopic properties of the mechanism that leads to it. Basic properties of the emerging structures (size distributions, longevity) are analyzed as well as their resilience against sensor or actuation noise. Finally, we explore parameter space for potential other candidates of life. The generality and simplicity of the motion law provokes the thought that one fundamental rule, described by one simple equation yields various structures in nature: it may work on different time- and size scales, ranging from the self-structuring universe, to emergence of living beings, down to the emergent subatomic formation of matter.

Lysophosphatidylcholine acts in the constitutive immune defence against American foulbrood in adult honeybees.

Honeybee (Apis mellifera) imagines are resistant to the Gram-positive bacterium Paenibacillus larvae (P. larvae), causative agent of American foulbrood (AFB), whereas honeybee larvae show susceptibility against this pathogen only during the first 48 h of their life. It is known that midgut homogenate of adult honeybees as well as a homogenate of aged larvae exhibit strong anti-P. larvae activity. A bioactivity-guided LC-HRMS analysis of midgut homogenate resulted in the identification of 1-oleoyl-sn-glycero-3-phosphocholine (LPC) pointing to a yet unknown immune defence in adult honeybees against P. larvae. Antimicrobial activity of LPC was also demonstrated against Melissococcus plutonius, causative agent of European Foulbrood. To demonstrate an AFB-preventive effect of LPC in larvae, artificially reared larvae were supplemented with LPC to evaluate its toxicity and to assess whether, after infection with P. larvae spores, LPC supplementation prevents AFB infection. 10 µg LPC per larva applied for 3 d significantly lowered mortality due to AFB in comparison to controls. A potential delivery route of LPC to the larvae in a colony via nurse bees was assessed through a tracking experiment using fluorescent-labelled LPC. This yet undescribed and non-proteinous defense of honeybees against P. larvae may offer new perspectives for a treatment of AFB without the utilization of classic antibiotics.

Effect of hydroxymethylfurfural (HMF) on mortality of artificially reared honey bee larvae (Apis mellifera carnica).

Hydroxymethylfurfural (HMF) is a heat-formed, acid-catalyzed contaminant of sugar syrups, which find their way into honey bee feeding. As HMF was noted to be toxic to adult honey bees, we investigated the toxicity of HMF towards larvae. Therefore we exposed artificially reared larvae to a chronic HMF intoxication over 6 days using 6 different concentrations (5, 50, 750, 5000, 7500 and 10,000 ppm) and a control. The mortality was assessed from day 2 to day 7 (d7) and on day 22 (d22). Concentrations ranging from 5 to 750 ppm HMF did not show any influence on larval or pupal mortality compared to controls (p > 0.05; Kaplan-Meier analysis). Concentrations of 7500 ppm or higher caused a larval mortality of 100%. An experimental LC50 of 4280 ppm (d7) and 2424 ppm (d22) was determined. The calculated LD50 was 778 µg HMF per larva on d7 and 441 µg HMF on d22. Additionally, we exposed adult honey bees to high concentrations of HMF to compare the mortality to the results from larvae. On d7 larvae are much more sensitive against HMF than adult honey bees after 6 days of feeding. However, on d22 after emergence adults show a lower LC50, which indicates a higher sensitivity than larvae. As toxicity of HMF against honey bees is a function of time and concentration, our results indicate that HMF in supplemental food will probably not cause great brood losses. Yet sublethal effects might decrease fitness of the colony.

Early maternal loss affects social integration of chimpanzees throughout their lifetime.

The long-term effects of early adverse experiences on later psychosocial functioning are well described in humans, but sparsely documented for chimpanzees. In our earlier studies, we investigated the effects of maternal and social deprivation on three groups of ex-laboratory chimpanzees who experienced either an early or later onset of long-term deprivation. Here we expand our research by adding data on subjects that came from two stable zoo groups. The groups comprised of early maternally deprived wild-caught chimpanzees and non-deprived zoo-born chimpanzees. We found that compared to zoo chimpanzees, ex-laboratory chimpanzees were more restricted regarding their association partners in the newly formed groups, but not during their second year of group-life, indicating that social stability has an important influence on the toleration of association partners close-by. Social grooming activity, however, was impaired in early long-term deprived ex-laboratory chimpanzees as well as in early maternally deprived zoo chimpanzees compared to non-deprived zoo chimpanzees. Thus, we conclude that early maternal loss has lifelong effects on the social integration of chimpanzees which becomes evident in their grooming networks. Although the retrospective nature of our study prevents a clear causal explanation, our results are of importance for understanding the development of social competence in chimpanzees.

From honeybees to robots and back: division of labour based on partitioning social inhibition.

In this paper, a distributed adaptive partitioning algorithm inspired by division of labor in honeybees is investigated for its applicability in a swarm of underwater robots in one hand and is qualitatively compared with the behavior of honeybee colonies on the other hand. The algorithm, partitioning social inhibition (PSI), is based on local interactions and uses a simple logic inspired from age-polyethism and task allocation in honeybee colonies. The algorithm is analyzed in simulation and is successfully applied here to partition a swarm of underwater robots into groups demonstrating its adaptivity to changes and applicability in real world systems. In a turn towards the inspiration origins of the algorithm, three honeybee colonies are then studied for age-polyethism behaviors and the results are contrasted with a simulated swarm running the PSI algorithm. Similar effects are detected in both the biological and simulated swarms suggesting biological plausibility of the mechanisms employed by the artificial system.

Immune responses of honeybees and their fitness costs as compared to bumblebees.

Immune responses of invertebrates imply more than developing a merely unspecific response to an infection. Great interest has been raised to unveil whether this investment into immunity also involves fitness costs associated to the individual or the group. Focusing on the immune responses of honeybees, we use the well-studied insect bumblebee for comparison. Bumblebees are capable of producing specific immune responses to infections whereas this has not been assessed for honeybees so far. We investigated whether a prior bacterial encounter provides protection against a later exposure to the same or a different bacterium in honeybees. Additionally, we studied whether the foraging activities of honeybees and bumblebees are affected upon immune stimulation by assessing the flight performance. Finally, the acceptance behavior of nestmates toward immune-challenged honeybees was determined. Results show that despite stimulating the immune system of honeybees, no protective effects to infections were found. Further, honeybees were not affected by an immune challenge in their flight performance whereas bumblebees showed significant flight impairment. Immune-challenged honeybees showed lower survival rates than naive individuals when introduced into a regular colony. Here, we reveal different immune response-cost scenarios in honeybees and bumblebees for the first time.