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International audience; To provide a complete portrayal of the multiple factors negatively impacting insects in agricultural landscapes it is necessary to assess the concurrent incidence, magnitude, and interactions among multiple stressors over substantial biogeographical scales. Trans-national ecological field investigations with wide-ranging stakeholders typically encounter numerous challenges during the design planning stages, not least that the scientific soundness of a spatially replicated study design must account for the substantial geographic and climatic variation among distant sites. ‘PoshBee’ (Pan-European assessment, monitoring, and mitigation of Stressors on the Health of Bees) is a multi-partner transdisciplinary agroecological project established to investigate the suite of stressors typically encountered by pollinating insects in European agricultural landscapes. To do this, PoshBee established a network of 128 study sites across eight European countries and collected over 50 measurements and samples relating to the nutritional, toxicological, pathogenic, and landscape components of the bees’ environment. This paper describes the development process, rationale, and end-result of each aspect of the of the PoshBee field investigation. We describe the main issues and challenges encountered during the design stages and highlight a number of actions or processes that may benefit other multi-partner research consortia planning similar large-scale studies. It was soon identified that in a multi-component study design process, the development of interaction and communication networks involving all collaborators and stakeholders requires considerable time and resources. It was also necessary at each planning stage to be mindful of the needs and objectives of all stakeholders and partners, and further challenges inevitably arose when practical limitations, such as time restrictions and labour constraints, were superimposed upon prototype study designs. To promote clarity for all stakeholders, for each sub-component of the study, there should be a clear record of the rationale and reasoning that outlines how the final design transpired, what compromises were made, and how the requirements of different stakeholders were accomplished. Ultimately, multi-national agroecological field studies such as PoshBee benefit greatly from the involvement of diverse stakeholders and partners, ranging from field ecologists, project managers, policy legislators, mathematical modelers, and farmer organisations. While the execution of the study highlighted the advantages and benefits of large-scale transdisciplinary projects, the long planning period emphasized the need to formally describe a design framework that could facilitate the design process of future multi-partner collaborations.

Honey is an important food for man and has been used as a natural sweetener since ancient times. It is a viscous and aromatic product made by honey bees using the nectar of flowers or honeydew. Hon...

The validation of a multiresidue method for the determination of organochlorine, organophosphorus, pyrethroid and dicarboximide pesticides in honeybees is described. The method involves the extraction of 25 pesticides using acetonitrile, liquid partitioning with n-hexane and a clean-up performed on a silica gel cartridge (1 g, 12 mL). Capillary gas chromatography with electron-capture and nitrogen–phosphorus detectors was used for analytes determination. Limit of quantification (LOQ), recovery and relative standard deviation (RSD) for each analyte were determined. The recovery data were obtained by spiking honeybee samples free of pesticides at three levels (LOQ, 4LOQ and 8LOQ) with organochlorine, organophosphorus, pyrethroid and dicarboximide pesticides. The recoveries were in the range between 90.0% and 101.5% for LOQ, between 90.3% and 104.8% for 4LOQ and between 88.1% and 101.6% for 8LOQ with RSD less than 20% for the three levels tested. The lowest LOQ value was 1 ng bee−1 (corresponding to ...

Co-exposure to pesticides and viruses is likely to occur in honey bee colonies. Pesticides can be present in pollen, nectar, and persist in stored food (honey and bee bread), and viruses can be highly prevalent in honey bee colonies. Therefore, the present study describes the influence of chronic co-exposure to thiamethoxam and Chronic bee paralysis virus (CBPV) on bee survival, virus loads, expression level of immune and detoxication genes, and pesticide metabolism Experiments were performed on honey bees collected from a winter apiary with reduced viral contaminations. No synergistic effect of co-exposure was observed on bee survival, nor on the ability of bees to metabolise the pesticide into clothianidin. However, we found that co-exposure caused an increase in CBPV loads that reached the viral levels usually found in overt infections. The effect of co-exposure on CBPV replication was associated with down-regulation of vitellogenin and dorsal-1a gene transcription. Nevertheless, the observed effects might be different to those occurring in spring or summer bees, which are more likelyco-exposed to thiamethoxam and CBPV and exhibit a different physiology.

The number of bees, amount of brood and haemolymph vitellogenin titre are parameters to establish the vitality of a honey bee colony. Increasing numbers of bees during summer until autumn; increasing amounts of brood in spring towards summer followed by a decrease; and low haemolymph vitellogenin titres in foraging bees and high vitellogenin titres in nurse bees and winter bees are typical courses over the bee season. In contrast to the number of bees and amount of brood, determined at the colony level, vitellogenin is determined in individual bees. To turn the factor vitellogenin from the mean of individual data into a colony parameter, a protocol was developed to determine the colony fraction vitellogenin in a pooled haemolymph sample of 30 bees, sampled from a defined location in the colony. This colony fraction vitellogenin is a parameter for honey bee colony vitality.

Pathogens and pesticides are likely to co-occur in honeybee hives, but much remains to be investigated regardingtheir potential interactions. Here, we first investigated the metabolisation kinetics of thiamethoxam in chronicallyfed honeybees. We show that thiamethoxam, at a dose of 0.25 ng/bee/day, is quickly and effectively metabolised into clothianidin, throughout a 20 day exposure period. Using a similar chronic exposure to pesticide, we then studied, in a separate experiment, the impact of thiamethoxam and Chronic bee paralysis virus (CBPV) co-exposure in honeybees. The honeybees were exposed to the virus by contact, mimicking the natural transmission route in the hive. We demonstrate that a high dose of thiamethoxam (5.0 ng/bee/day) can cause a synergistic increase in mortality in co-exposed honeybees after 8 to 10 days of exposure, with no increase in viral loads. At a lower dose (2.5 ng/bee/day), there was no synergistic increase of mortality, but viral loads were significantly higher in naturally dead honeybees, compared with sacrificed honeybees exposed to the same conditions. These results show that the interactions between pathogens and pesticides in honeybees can be complex: increasing pesticide doses may not necessarily be linked to a rise in viral loads, suggesting that honeybee tolerance to the viral infection might change with pesticide exposure.

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The validation of a multi-residue method for the determination of five neonicotinoid insecticides (imidacloprid, clothianidin, acetamiprid, thiacloprid and thiamethoxam) in honeybees is described. The method involves the extraction of pesticides using acetonitrile and liquid partitioning with n-hexane. One clean-up is then performed on a florisil cartridge (1 g, 6 mL) and the extract is analysed by liquid chromatography-electrospray ionisation-tandem mass spectrometry (LC-ESI-MS/MS). The recovery data were obtained by spiking honeybees samples free of pesticides at two concentration levels of the various neonicotinoids. The recoveries were in the range between 93.3 and 104.0% with relative standard deviation (RSD) less than 20%. The limit of quantification (LOQ) was 0.5 ng g−1 (corresponding to 0.05 ng bee−1) for all pesticides except for acetamiprid which was 1 ng g−1 (corresponding to 0.1 ng bee−1).

SummarySeveral cases of mortality of honey bee colonies (varying from 38 to 100%) were observed in France during the winter of 2005–6. In order to explain the causes of these mortalities, a case control study was conducted on a limited area, together with a larger survey in 18 other apiaries located in 13 sites over the entire country. Both studies included diagnosis of the main honey bee diseases, assessment of the colony management measures taken by beekeepers and the determination of pesticide residues in apicultural matrices. Pollen analysis was carried out on beebread samples to identify which floral species were used for forage before colony death. Poor Varroa destructor treatments together with Nosema disease and brood diseases were frequent in apiaries with high colony mortalities. The absence of any preventive treatment against V. destructorwas the main risk factor.

International audience; A 3-yr field survey was carried out in France, from 2002 to 2005, to study honey bee (Apis mellifera L.) colony health in relation to pesticide residues found in the colonies. This study was motivated by recent massive losses of honey bee colonies, and our objective was to examine the possible relationship between low levels of pesticide residues in apicultural matrices (honey, pollen collected by honey bees, beeswax) and colony health as measured by colony mortality and adult and brood population abundance. When all apicultural matrices were pooled together, the number of pesticide residue detected per sampling period (four sampling periods per year) and per apiary ranged from 0 to 9, with the most frequent being two (29.6%). No pesticide residues were detected during 12.7% of the sampling periods. Residues of imidacloprid and 6- chloronicotinic acid were the most frequently detected in pollen loads, honey, and honey bee matrices. Several pairs of active ingredients were present concurrently within honey bees and in pollen loads but not in beeswax and honey samples. No statistical relationship was found between colony mortality and pesticide residues. When pesticide residues from all matrices were pooled together, a mixed model analysis did not show a significant relationship between the presence of pesticide residues and the abundance of brood and adults, and no statistical relationship was found between colony mortality and pesticide residues. Thus, although certain pesticide residues were detected in apicultural matrices and occasionally with another pesticide residual, more work is needed to determine the role these residues play in affecting colony health.

Acaricide residues were assessed in French commercial beeswax using newly developed liquid and gas chromatography methods. Most of the commercial wax samples and all samples taken during the industrial recycling process contained coumaphos and fluvalinate. Amitraz and coumaphos residue levels were also followed in several hives experimentally treated with Asuntol®50 or Apivar®, two products used in France to control varroa infestation. After the Asuntol®50 treatment, coumaphos residues increased in honey and wax combs, persisted more than 30 days in honey and one year or more in comb wax. The half-life of coumaphos was 69 and 115–346 days in honey and comb wax respectively. Following Apivar® treatment, amitraz was not detected in honey nor in wax. These results are consistent with and complete other studies: the use of coumaphos entails wax contamination which persists through commercial recycling. As this may be a threat for bee health, the use of Asuntol®50 should be avoided.

In 2002, a field survey was initiated on French apiaries to monitor weakness of honey bee, Apis mellifera L., colonies. Apiaries were evenly distributed in five sites located on continental France. Five colonies were randomly selected in each apiary, leading to a total of 125 studied honey bee colonies. For 3 yr (starting in autumn 2002), colonies were visited four times per year: after winter, before summer, during summer, and before winter. Pollen loads from traps were collected at each visit. Multiresidue analyses were performed in pollen to search residues of 36 different molecules. Specific analyses were conducted to search fipronil and metabolites and also imidacloprid and metabolites. Residues of 19 searched compounds were found in samples. Contamination by pesticides ranged from 50 to 0%. Coumaphos and tau-fluvalinate residues were the most concentrated of all residues (mean concentrations were 925.0 and 487.2 microg/kg, respectively). Fipronil and metabolite contents were superior to the limit of detection in 16 samples. Residues of fipronil were found in 10 samples. Nine samples contained the sulfone compound, and three samples contained the desulfinyl compound. Residues of imidacloprid and 6-chloronicotinic acid were found in 69% of samples. Imidacloprid contents were quantified in 11 samples with values ranging from 1.1 to 5.7 microg/kg. 6-Chloronicotinic acid content was superior to the limit of quantification in 28 samples with values ranging from 0.6 to 9.3 microg/kg. Statistical tests showed no difference between places of sampling with the exception of fipronil. Possible origins of these contaminations, concentration and toxicity of pesticides, and the possible consequences for bees are discussed.

Two groups of eight honey bee colonies were fed with two different concentrations of imidacloprid in saccharose syrup during summer (each colony was given 1 litre of saccharose syrup containing 0.5 microg litre(-1) or 5 microg litre(-1) of imidacloprid on 13 occasions). Their development and survival were followed in parallel with control hives (unfed or fed with saccharose syrup) until the end of the following winter. The parameters followed were: adult bee activity (number of bee entering the hive and pollen carrying activity), adult bee population level, capped brood area, frequency of parasitic and other diseases, mortality, number of frames with brood after wintering and a global score of colonies after wintering. The only parameters linked to feeding with imidacloprid-supplemented saccharose syrup when compared with feeding with non-supplemented syrup were: a statistically non-significant higher activity index of adult bees, a significantly higher frequency of pollen carrying during the feeding period and a larger number of capped brood cells. When imidacloprid was no longer applied, activity and pollen carrying were re-established at a similar level for all groups. Repeated feeding with syrup supplemented with imidacloprid did not provoke any immediate or any delayed mortality before, during or following the next winter, whereas such severe effects are described by several French bee keepers as a consequence of imidacloprid use for seed dressing in neighbouring cultures. In any case, during the whole study, mortality was very low in all groups, with no difference between imidacloprid-fed and control colonies. Further research should now address several hypotheses: the troubles described by bee keepers have causes other than imidacloprid; if such troubles are really due to this insecticide, they may only be observed either when bees consume contaminated pollen, when no other sources of food are available, in the presence of synergic factors (that still need to be identified), with some particular races of bees or when colonies are not strong and healthy.

A rapid reversed‐phase high‐performance liquid chromatographic method, with fluorimetric detection, is described for routine analysis of sulfathiazole (STZ) residues in honey. Samples were dissolved in 1 M hydrochloric acid and submitted to pre‐column derivatization with fluorescamine. Liquid chromatographic analysis of the fluorescent derivative is performed on a C18 column using a mobile phase of 2% acetic acid/ acetonitrile (60:40, v/v). The quantification limit obtained for STZ in honey was 10 µg/kg.

Rapid analytical methods are described to control quality of honeys, concerning residues of acaricides applied in hives to prevent Varroa jacobsoni infestation. A liquid-liquid extraction with hexane-propanol-2-ammonia (60 ml:30 ml:0.28%) was used for the simultaneous analysis of coumaphos, bromopropylate, amitraz and fluvalinate. For thymol, one clean up on a solid-phase extraction C18 (500 mg, 6 ml) column was performed; for rotenone, a liquid extraction with dichloromethane was realised. Quantitative recoveries obtained with honey were satisfactory and were superior to 80%. All acaricides are identified by reversed-phase high-performance liquid chromatography with photodiode array detection. Quantification limits obtained were below maximal residue limits when these exist.

(2010). A case report of a honey bee colony poisoning incident in France. Journal of Apicultural Research: Vol. 49, Honey bee colony losses, pp. 113-115.

The determination of iprodione, vinclozolin, and cymoxanil fungicide residues in raspberries and lettuce has been carried out by HPLC with an UV detector, by HPTLC with an densitometric detection, and by GC with an electron capture detector (ECD). In all cases, fungicide residues were extracted with acetone and liquid-liquid partitioning process and finally purified on silica gel. Recoveries are always higher than 70%. For iprodione, vinclozolin, and cymoxanil, respectively, the limits of detection were 0.01 ppm, 0.013 ppm, and 0.08 ppm for the HPLC method, 0.2 ppm, 0.43 ppm, and 0.5 ppm for the HPTLC method, and 0.025 ppm, 0.004 ppm, and 0.03 ppm for the GC method.

The frequency of occurrence and relative concentration of 44 pesticides in apicultural (Apis mellifera) matrices collected from five French locations (24 apiaries) were assessed from 2002 to 2005. The number and nature of the pesticides investigated varied with the matrices examined-living honeybees, pollen loads, honey, and beeswax. Pollen loads and beeswax had the highest frequency of pesticide occurrence among the apiary matrices examined in the present study, whereas honey samples had the lowest. The imidacloprid group and the fipronil group were detected in sufficient amounts in all matrices to allow statistical comparisons. Some seasonal variation was shown when residues were identified in pollen loads. Given the results (highest frequency of presence) and practical aspects (easy to collect; matrix with no turnover, unlike with bees that are naturally renewed), pollen loads were the best matrix for assessing the presence of pesticide residues in the environment in our given conditions.

International audience; Tetracyclines are used to control bacterial diseases like European and American foulbrood which may cause severe losses in honey bee population and honey production. This study, using 24 hives randomly distributed into four groups of 6 hives, was performed for measuring the occurrence of tetracycline (TC) residue levels in honey following two types of TC application. Two groups of colonies were treated 3 times with 0.5 g of TC in 1 litre of syrup (group S) or in 10 g of powdered sugar (group P). Six hives of a first control group (group C) fed with untreated syrup were installed at 20 and 45 metres from groups S and P respectively. A second control group (group DC) was disposed 3 km apart. TC residues in honey sampled at different times in all hives and in honey artificially contaminated with TC and stored in the laboratory at 4, 20 and 35°C were analysed by ELISA and HPLC methods. One day after the last application, the mean TC concentration in brood chamber honey was ten times higher in group S (40.7 mg.kg-1) than in group P (4.34 mg.kg-1). After 8 days, TC residues were detected in all hives of group C. After 146 days, the mean TC concentration in harvested honey was 1.54, 0.35, and 0.15 mg.kg-1 for groups S, P and C respectively. The control group C had been contaminated by drifting. In all hives of the group DC, no residues were detected at any time of the study. The honey harvested at day 504 did not contained any detectable TC residues, except in one super from group C (0.026 mg.kg-1). The half-life time of TC in honey from supers was similar in groups C, S and P and equal to 65 days. This duration was twice lower than in honey stored in laboratory in similar conditions : at 35°C in the dark (t1/2 = 121 days). In honey stored at 20°C, TC was quite stable as its half-life time was 242 days. The data from these experiments specify levels of TC residues in honey after a treatment in hives, their persistence and diffusion into the apiary. These results show that the TC must be used with precaution in honey production.

Two groups of eight honey bee colonies were fed with two different concentrations of imidacloprid in saccharose syrup during summer (each colony was given 1 litre of saccharose syrup containing 0.5 microg litre(-1) or 5 microg litre(-1) of imidacloprid on 13 occasions). Their development and survival were followed in parallel with control hives (unfed or fed with saccharose syrup) until the end of the following winter. The parameters followed were: adult bee activity (number of bee entering the hive and pollen carrying activity), adult bee population level, capped brood area, frequency of parasitic and other diseases, mortality, number of frames with brood after wintering and a global score of colonies after wintering. The only parameters linked to feeding with imidacloprid-supplemented saccharose syrup when compared with feeding with non-supplemented syrup were: a statistically non-significant higher activity index of adult bees, a significantly higher frequency of pollen carrying during the feeding period and a larger number of capped brood cells. When imidacloprid was no longer applied, activity and pollen carrying were re-established at a similar level for all groups. Repeated feeding with syrup supplemented with imidacloprid did not provoke any immediate or any delayed mortality before, during or following the next winter, whereas such severe effects are described by several French bee keepers as a consequence of imidacloprid use for seed dressing in neighbouring cultures. In any case, during the whole study, mortality was very low in all groups, with no difference between imidacloprid-fed and control colonies. Further research should now address several hypotheses: the troubles described by bee keepers have causes other than imidacloprid; if such troubles are really due to this insecticide, they may only be observed either when bees consume contaminated pollen, when no other sources of food are available, in the presence of synergic factors (that still need to be identified), with some particular races of bees or when colonies are not strong and healthy.

International audience; This review covers mainly publications that appeared in Analytical Abstracts (Royal Society of Chemistry) from January 1990 to February 2001. The number of publications on this topic continues to grow, and during the past three years (1998-2000) about 150 reviews and/or overviews have been published in the area of food. Numerous techniques and food matrices or chemical components are presented and discussed in these reviews. The present review is intentionally limited to eight techniques or classes of techniques and intends to be a "technique by technique" presentation of "what was used" or "what is used" to characterize food products and to detect their possible adulteration. The present review focuses on the following techniques: microscopic analysis; HPLC; GC, GC-(MS, FTIR); UV-visible spectrophotometry; AAS/AES, ICP-(AES, MS); IRMS, GC-IRMS, GC-C-IRMS; DSC; IR, mid-IR, and NMR (202 references). Emphasis is placed as much as possible on chemometrical treatment of analytical data, which are commonly used to achieve the final objective, either food characterization or adulteration detection. Finally, a brief description is given of the new generation of analytical systems that combine powerful analytical techniques and powerful computer software for a best extraction of the information from analytical data.

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