Effect of Apis mellifera on community composition of local pollinator bees and their pollination network in Qinling Mountains and surrounding areas

The Qinling Mountains, known for their rich vegetation and diverse pollinating insects, have seen a significant decline in bee species richness and abundance over recent decades, largely due to the introduction and spread of Apis mellifera. This decline has caused cascading effects on the region's community structure and ecosystem stability. To improve the protection of native bees in the natural and agricultural landscape of the Qinling Mountains and its surrounding areas, we investigated 33 sampling sites within three habitats: forest, forest-agriculture ecotones, and farmland. Using a generalized linear mixing model, t-test, and other data analysis methods, we explored the impact of Apis mellifera on local pollinator bee richness, abundance, and the pollination network in different habitats in these regional areas. The results show that(1)Apis mellifera significantly negatively affects the abundance and richness of wild pollinator bees,while Apis cerana abundance is also affected by beekeeping conditions.(2)There are significant negative effects of Apis mellifera on the community structure of pollinator bees in the Qinling Mountains and its surrounding areas: the Shannon-Wiener diversity index, Pielou evenness index, and Margalef richness index of bee communities at sites with Apis mellifera influence were significantly lower than those at sites without Apis mellifera influence.(3)The underlying driver of this effect is the monopolization of flowering resources by Apis mellifera. This species tends to visit flowering plants with large nectar sources, which constitute a significant portion of the local plant community. By maintaining a dominant role in the bee-plant pollination network, Apis mellifera competitively displaces native pollinator bees, reducing their access to floral resources. This ultimately leads to a reduction in local bee-plant interactions, decreasing the complexity and stability of the pollination network. These findings highlight the need for targeted conservation efforts to protect native pollinator species and maintain the ecological balance in the Qinling Mountains.

Further Studies of the Cap Pushing Response in Honey Bees (Apis mellifera)

The Cap Pushing Response (CPR) is a free-flying technique used to study learning and memory in honey bees (Apis mellifera). The series of experiments outlined in this paper aimed to test whether honey bees exhibit the cognitive concept of “expectancy” utilizing the CPR in a weight differentiation paradigm. Five previous experiments in our laboratory have explored whether the concept of expectancy can account for honey bee performance and have all failed to support the cognitive interpretation. The first experiment examined if bees could differentiate between the two caps in the amount of force they used to push the cap and the distance the cap was pushed when the caps were presented one at a time. The second experiment explored cap weight preference by presenting bees with a choice between the two caps. The third and fourth experiments tested the bee’s ability to expect reward or punishment based on cap weight. Results revealed that bees were found to have a strong preference for the light cap and therefore were not able to expect reward or punishment based on cap weight. These experiments contribute to the debate on whether bees have “cognitive” representations and continue to support the behaviorist interpretation.

Correlation between the Change in the Number of Apis mellifera Worker Bees and Nectar Secretion of Nectariferous Plants

[Objective] This study aimed to clarify the correlation between changes of Apis mel ifera and the nectar secretion characteristics of nectariferous plants. [Method] Considering the nectar secretion characteristics of major and auxiliary nec-tariferous plants, six Apis mel ifera colonies were selected for measure the number of eggs, larvae, pupae and adult bees from Jan. to Dec. in 2012; based on that, their annual change curves were also plotted. [Result] The results showed that there were three peaks of the total number of A. mel ifera workers throughout the year：the first occurred on May 15th, with bees developed into an ideal population for col-lecting pomegranate nectar, and the second and third peaks occurred on July 15th and Oct. 15th, respectively, with bees developed into an ideal population for col ect-ing E. ciliate （Thuab） Hyland. [Conclusion] Prevention of Varroa jacobsoni should be carried out with two or more types of acaricides at the late nectar flow stages of the two nectariferous plants（pomegranate and E. ciliate （Thuab） Hyland） when there was a nectar deficiency. Prevention of Tropilaelaps clareae should be timely per-formed with sublimed sulfur in conjunction with acaricides. This study provides a theoretical basis for the high-quality and high-yielding production of honey, as wel as for the product safety.

cDNA Cloning and Bioinformatic Prediction of TPI Gene from Apis mellifera

[Objective] The aim was to clone triosephosphate isomerase （TPI） gene from Apis mellifera, and predict the properties of TPI protein with bioinformatic meth- ods. [Method] The TPI gene was firstly cloned by in silico cloning based on the ex- pressed sequence tags （ESTs） from Unigene of NCBI. Some characters of the TPI protein including hydrophobicity or hydrophilicity, isoelectric point （pl） and secondary structure were analyzed and predicted by the tools of bioinformatics. [Result] The TPI gene from A. mellifera was 1 768 bp in full length and it contained a complete ORF which encoded 247 amino acids; the pl of TPI protein was 8.515; the TPI protein was a member of ~13-fold family. [Conclusion] The in silico cloning based on the expressed sequence tags is a efficient method in practice, and this study will provide more references for further study on A. mellifera at molecular level.

Foraging and Pollination Behaviour of Apis mellifera adansonii Latreille （Hymenoptera, Apidae） on Gossypium hirsutum （Malvaceae） Flowers at Dang （Ngaoundere, Cameroon）

To evaluate Apis mellifera adansonii Latreille impact on fruit and seed yields of Gossypium hirsutum L., its foraging and pollinating activities were studied in Ngaoundere for two seasons. Observations were made on 340 flowers each year and divided in three treatments. The treatments included unlimited flowers access by all visitors; bagged flowers to deny all visits and limited visits by Apis mellifera adansonii only. The worker bees seasonal rhythm of activity, its foraging behaviour, its pollination efficiency, the fruiting rate, the number of seeds per fruit and the percentage of normal seeds were evaluated. Results show that this bee foraged G. hirsutum flowers throughout the whole blooming period. This bee species intensely harvested pollen and nectar. The mean foraging speed was 9.41 flowers per rain in 2009 and 8.41 flowers per min in 2010. The fruiting rate, the number of seeds per fruit and the percentage of normal seeds of unprotected flowers were significantly higher than those of flowers protected from insects. Through its pollination efficiency, Apis mellifera adansonii provoked a significant increment of the fruiting rate by 60.84% in 2009 and 36.48% in 2010, as well as the number of seeds per fruit by 94.16% in 2009 and 31.41% in 2010, and the percentage of normal seeds by 94.23% in 2009 and 33.49% in 2010. The installation ofA. m. adansonii colonies close to G. hirsutum fields could be recommended to increase fruit, seed and honey yields, and pollen production as a hive product in the region.

The Formamidine Amitraz as a Hyperglycemic α-Agonist in Worker Honeybees(Apis mellifera mellifera L.)in Vivo

Intra-abdominal injection of amitraz(0.25 nmol per honeybee,i.e.,approx 2.3 nmol/g)to emerging worker bees,in vivo,led to a significant hypertrehalosemia(300-400%)followed by a hyperglucosemia(≈600%).Maxima were reached at 0.5 and 2h,respectively.A strong negative correlation between glucosemia and trehalosemia appeared after injection of pure phentolamine (1 nmol per bee),suggesting stimulation of trehalase activities.Simultaneous administration of the α-blocker at≥0.25 nmol per individual suppressed the hyperglycemic response of amitraz. The formamidine pesticide thus likely acts on the honeybee α-aminergic system.1989 Academic Press,Inc.

Cloning and Sequence Analysis of Carbonic Anhydrase-related Protein 10-like in Apis mellifera

[ Objective ] This study aimed to clone and analyze the gene sequence encoding carbonic anhydrase-related protein lO-like （ CARP X） from Apis mellif era. [Method] The cDNA sequence of CARPX gene was cloned through RT-PCR, and then analyzed with bioinformatic method. [Result] The full-length cDNA sequence of CARPX was 972 bp long and encoded 324 amino acid residues, including a signal peptide and two transmembrane domains. The predicted molecular mass was 37.1 kDa and the predicted isoeleetric point was 7.458. The CARP X from A. mellifera shared close relationship with proteins from Apisflorae, Bombtas impatiens, Bombus terrestris, Nasonia vitripennis and Acyrthosiphon pisum. The insect CARP X family may include two subfamilies. [ Conclusion] The results pro- vide basis for studying CARPs family.

Genetic Variability of the Mitochondrial DNA in Honeybees （Apis mellifera L.） from Benin

The aim of this study was to evaluate the genetic variability in bees Apis mellifera from Benin by using mitochondrial DNA （mtDNA） as a molecular marker in their cytochrome c oxidase subunit I and II （COI-COI1） intergenic region. A total of 304 bee colonies were sampled in 27 municipalities of the cashew growing area of Benin. These samples were analyzed by the cleaved amplified polymorphisms technique for determining the haplotypes of subspecies present in the sampled population. Eight PCR-RFLP profiles of African lineage A were then identified in the 304 samples of bees investigated. Forty-nine percent （49%） of the samples showed the profile of haplotype A1 （subspecies adansonii of Zambia）, 40% of haplotype A4 （subspecies scutellata of South Africa） and 3% of haplotype A 19 （subspecies adansonii of Guinea）. Five other haplotypes of the African branch （A） that had been described in a previous study were also identified： new 1 （2%）, new 2 （2%）, new 3 （1%）, new 4 （2%） and new 5 （1%）. This study showed that A. rnellifera from Benin belonged only to lineage A with the predominance of haplotypes AI and A4. This study will contribute to the development of coherent policies for conservation of local bees in Benin.

Evaluation of Morphometric Characters of Honeybee (Apis mellifera L.) Populations in the Lake Chad Basin in Central Africa

Samples of workers of honeybee were collected from 42 colonies in 13 localities in the Lake Chad Basin (parts of Cameroon, Chad, and Nigeria) and analysed using classical morphometry. Measurements of 35 morphological characters of body size, colour and pilosity were taken from 10 workers per colony and the data subjected to one-way analysis of variance (ANOVA), principal components analysis (PCA), hierarchical cluster analysis (HCA), stepwise discriminant analysis (DA) and Pearson’s product-moment correlation analysis. A one-way ANOVA revealed that means of 21 of the morphometric characters differed significantly (p 0.05). The bees formed one cluster in a PCA. However, scatter plots of altitude against principal component 1 of PCA (loaded with characters of body size) revealed an increase of size of the bees along the gradient of the Lake Chad Basin. The coefficient of determination (R2) indicated that 88% and 77% of the variation in size might be explained by altitude in the southeastern and southwestern parts of the basin, respectively. Additionally, there was a very highly significant strong positive relationship between principal component 1 and altitude (r(30) = 0.618, p < 0.0005). Similarly, HCA and DA classified the colonies into three morphoclusters whose distribution closely followed the altitude of the area.

Surface Temperature and Heat Transfer between Body Regions of Africanized Honeybees(Apis mellifera L.)in Hives under Sun and Shade Conditions in the Northeastern Semi-arid Region of Brazil

This work was done at the experimental station of the Federal Rural University of the Semi-arid Region （UFERSA） in Mossoró, Brazil. This work aimed to evaluate the surface temperature of the bees and heat transfer between body regions of Africanized honeybees （Apis mellifera L.） in hives under sun and shade conditions in the northeastern semi-arid region of Brazil. Twelve colonies of Africanized honeybees housed in Langstroth hives were used. The body surface temperature of the bees was measured using a mini infrared thermometer （Fluke model 62）. The climatological data were obtained through a meteorological station. The comparison of means was performed by the Tukey-Kramer test （p 〈 0.01）, and data analysis using the Statistical Analysis System software. The body temperatures of the bees are not homogeneous and fluctuate throughout the day, in accordance with the environmental variations. The temperature of the thorax is generally higher, followed by the head and abdomen. The head and the abdomen are parts of the body for which the excess temperature of the thorax is transferred, with the head being the main one. The bees that were in the shade, managed to keep their body temperatures at relatively normal levels, while the bees that were in the sun, had a considered increase of its surface body temperatures higher than 2 °C, being necessary the activation of mechanisms of heat loss, and consequently there is a deviation of energy that could be used for other activities within the colony.

Daily number of bee louse （Braula coeca） in honey bee （Apis mellifera carnica and A. m. syriaca） colonies maintained under semi-arid conditions

Experimental work was conducted at two apiaries located in Irbid district and in Shuna North, Jordan, during the years 2004-2006. The aims of these investigations were to estimate the seasonal changes in the infestation rates of the bee louse （Braula sp.） and to develop an easy and rapid method of estimating the infestation rate on workers with bee Braula. Two major honey bee subspecies are reared in Jordan; Apis mellifera carnica and Apis mellifera syriaca were used in this study. The results showed that the infestation rate began to increase rapidly in May, reaching the season＇s maximum rate of 16.2%, 15.8% and 17.4% forA. m. carnica and 22.6%, 23.9% and 22.9% forA. m. syriaca in December of 2004, 2005 and 2006, respectively. The maximum adult numbers of bees were found in April and June, whereas the minimum for the year was in January in both honey bee subspecies colonies during the study period. The actual population of the bee louse could be estimated by counting the daily dropped lice and multiplying by a factor of 158. This factor is valid for the experimental colonies of both subspecies kept for 3 years under semi-arid Mediterranean conditions.

The integrative analysis of microRNA and mRNA expression in Apis mellifera following maze-based visual pattern learning

The honeybee （Apis mellifera） is a social insect with strong sensory capacity and diverse behavioral repertoire and is recognized as a good model organism for studying the neurobiological basis of learning and memory. In this study, we analyzed the changes in microRNA （miRNA） and messenger RNA （mRNA） following maze-based visual learning using next-generation small RNA sequencing and Solexa/lllumina Digital Gene Expression tag profiling （DGE）. For small RNA sequencing, we obtained 13 367 770 and 13 132 655 clean tags from the maze and control groups, respectively. A total of 40 differentially expressed known miRNAs were detected between these two samples, and all of them were up-regulated in the maze group compared to the control group. For DGE, 5 681 320 and 5 939 855 clean tags were detected from the maze and control groups, respectively. There were a total of 388 differentially expressed genes between these two samples, with 45 genes up-regulated and 343 genes down-regulated in the maze group, compared to the control group. Additionally, the expression levels of 10 differentially expressed genes were confirmed by quantitative reverse transcription polymerase chain reaction （qRT-PCR） and the expression trends of eight of them were consistent with the DGE result, although the degree of change was lower in amplitude. The integrative analysis of miRNA and mRNA expression showed that, among the 40 differentially expressed known miRNAs and 388 differentially expressed genes, 60 pairs of miRNA/mRNA were identified as co-expressed in our present study. These results suggest that both miRNA and mRNA may play a pivotal role in the process of learning and memory in honeybees. Our sequencing data provide comprehensive miRNA and gene expression information for maze-based visual learning, which will facilitate understanding of the molecular mechanisms of honeybee learning and memory.

Next generation sequencing of Apis mellifera syriaca identifies genes for Varroa resistance and beneficial bee keeping traits

Apis mellifera syriaca exhibits a high degree of tolerance to pests and pathogens including varroa mites. This native honey bee subspecies of Jordan expresses behavioral adaptations to high temperature and dry seasons typical of the region. However, persistent honey bee imports of commercial breeder lines are endangering local honey bee population. This study reports the use of next-generation sequencing （NGS） technology to study the A. m. syriaca genome and to identify genetic factors possibly contributing toward mite resistance and other favorable traits. We obtained a total of 46.2 million raw reads by applying the NGS to sequence A. m. syriaca and used extensive bioinformatics approach to identify several candidate genes for Varroa mite resistance, behavioral and immune responses char- acteristic for these bees. As a part of characterizing the functional regulation of molecular genetic pathway, we have mapped the pathway genes potentially involved using information from Drosophila melanogaster and present possible functional changes implicated in responses to Varroa destructor mite infestation toward this. We performed in-depth functional annotation methods to identify -600 candidates that are relevant, genes involved in pathways such as microbial recognition and phagocytosis, peptidoglycan recognition protein family, Gram negative binding protein family, phagocytosis receptors, serpins, Toll signaling pathway, Imd pathway, Tnf, JAK-STAT and MAPK pathway, heamatopioesis and cellular response pathways, antiviral, RNAi pathway, stress factors, etc. were selected. Finally, we have cataloged function-specific polymorphisms between A. mellifera and A. m. syriaca that could give better understanding of varroa mite resistance mechanisms and assist in breeding. We have identified immune related embryonic development （Cactus, Relish, dorsal, Ank2, baz）, Varroa hygiene （NorpA2, Zasp, LanA, gasp, impl3） and Varroa resistance （Pug, pcmt, elk, elf3-s10, Dscam2, Dhc64C, gro, futsch） functional variations genes between A. mellifera and A. m. syriaca that could be used to develop an effective molecular tool for bee conservation and breeding programs to improve locally adapted strains such as syriaca and utilize their advantageous traits for the benefit of apiculture industry.

Genomics and proteomics of Apis mellifera filamentous virus isolated from honeybees in China

Apis mellifera filamentous virus(Am FV)is a large DNA virus that is endemic in honeybee colonies.The genome sequence of the Am FV Swiss isolate(Am FV CH–C05)has been reported,but so far very few molecular studies have been conducted on this virus.In this study,we isolated and purified Am FV(Am FV CN)from Chinese honeybee(Apis mellifera)colonies and elucidated its genomics and proteomics.Electron microscopy showed ovoid purified virions with dimensions of 300–500×210–285 nm,wrapping a 3165×40 nm filamentous nucleocapsid in three figure-eight loops.Unlike Am FV CH–C05,which was reported to have a circular genome,our data suggest that Am FV CN has a linear genome of approximately 493 kb.A total of 197 ORFs were identified,among which36 putative genes including 18 baculoviral homologs were annotated.The overall nucleotide similarity between the CN and CH–C05 isolates was 96.9%.Several ORFs were newly annotated in Am FV CN,including homologs of per os infectivity factor 4(PIF4)and a putative integrase.Phylogenomic analysis placed Am FVs on a separate branch within the newly proposed virus class Naldaviricetes.Proteomic analysis revealed 47 Am FV virionassociated proteins,of which 14 had over 50%sequence coverage,suggesting that they are likely to be main structural proteins.In addition,all six of the annotated PIFs(PIF-0–5)were identified by proteomics,suggesting that they may function as entry factors in Am FV infection.This study provides fundamental information regarding the molecular biology of Am FV.

Distribution and variability of deformed wing virus of honeybees （Apis mellifera） in the Middle East and North Africa

Three hundred and eleven honeybee samples from 12 countries in the Middle East and North Africa （MENA） （Jordan, Lebanon, Syria, Iraq, Egypt, Libya, Tunisia, Algeria, Morocco, Yemen, Palestine, and Sudan） were analyzed for the presence of deformed wing virus （DWV）. The prevalence of DWV throughout the MENA region was pervasive, but variable. The highest prevalence was found in Lebanon and Syria, with prevalence dropping in Palestine, Jordan, and Egypt before increasing slightly moving westwards to Algeria and Morocco Phylogenetic analysis of a 194 nucleotide section of the DWV Lp gene did not identify any significant phylogenetic resolution among the samples, although the sequences did show consistent regional clustering, including an interesting geographic gradient from Morocco through North Africa to Jordan and Syria. The sequences revealed several clear variability hotspots in the deduced amino acid sequence, which furthermore showed some patterns of regional identity. Furthermore, the sequence variants from the Middle East and North Africa appear more numerous and diverse than those from Europe.

Methionine as a methyl donor regulates caste differentiation in the European honey bee(Apis mellifera)

Nutrition contributes to honey bee caste differentiation,but the role of individual nutrients is still unclear.Most essential amino acid contents,except that of methionine(Met),are greater in royal jelly than worker jelly.After〜3.5 i the Met content in the latter was slightly greater than in the former.Met is the major raw material used in the synthesis of S-adenosyl-L-methionine,an active methyl donor for DNA methylation,which is an epigenetic driver of caste differentiation.Here,we tested whether Met regulates caste differentiation in honey bees by determining its effects on the caste development of bees receiving four diets:the basic,basic+0.2%Met,basic+0.2%Met+20 mg/kg 5-azacytidine,and basic+20 mg/kg 5-azacytidine.The presence of Met decreased the adult bee body length and the numbers of ovarioles,indicating that Met may direct the development of female larvae toward worker bees.The upregulated expression of SAMS,Dnmtl,and Dnmt3 caused by Met exposure in 4-d-old larvae indicated that the worker-inductive effects of Met may occur through the promotion of DNA methylation.We investigated the co-effects of Met and glucose on bee development,and found that the effects of an increased glucose level on the number of ovarioles and body length did not strengthen the worker-inductive effects caused by Met.Our results contribute to caste development theory and suggest that Met-as a methyl donor一plays a regulatory,but not decisive,role in caste differentiation.

Comparative Analyses of Proteome Complement Between Worker Bee Larvae of High Royal Jelly Producing Bees (A. m. ligustica) and Carniolian Bees (A. m. carnica)

This study is to compare the protein composition of the high royal jelly producing bee （A. m. ligustica） with that of Carniolian bee （A. m. carnica） during their worker larval developmental stage. The experiment was carried out by two- dimensional gel electrophoresis. The results showed that significant higher numbers of total proteins （283） were detected in larvae of high royal jelly producing bees （Jelly bee） than those of Carniolian bees （152） on 2-d-old larvae. Among them, 110 proteins were presented on both strains of bee larvae, whereas 173 proteins were specific to larvae of Jelly bees, and 42 proteins were exclusive to Carniolian larvae. However, on the 4th d, a significant higher number of total proteins （290） were detected in larvae of Jelly bees than those of Carniolian bees （240）, 163 proteins resolved to both bee larvae, and 127 proteins were specific to Jelly bees and 77 proteins to Camiolian bees. Until the 6th d, also a significant higher number of total proteins （236） were detected in larvae of Jelly bees than those of Carniolian bees （180）, 132 proteins were constantly expressed in two bee larvae, whereas 104 and 48 proteins are unique to Jelly bee and Carniolian bee larvae, respectively. We tentatively concluded that the metabolic rate and gene expression of Jelly bees larvae is higher than those of Carniolian bees based proteins detected as total proteins and proteins specific to each stage of two strains of bee larvae. Proteins constantly expressed on 3 stages of larval development with some significant differences between two bee strains, and proteins unique to each stage expressed differences in term of quality and quantity, indicating that larval development needed house keeping and specific proteins to regulate its growth at different development phage, but the expression mold is different between two strains of larval development.

The Effects of Temperature and Humidity around the Beehives on the Distribution of Nosema ceranae, and also Geographical and Seasonal Activity of the Infection in the Eastern Black Sea Region of Turkey.

20 localities were randomly selected in Eastern Black Sea Region of Turkey and samples were collected from around the beehives from April to September. Total of 4,640 dead adult worker bees were examined during the study. Total infection rate in worker bees was 21.23%. Nosema ceranae was identified in all localities with molecular techniques. Temperature and humidity values were measured from around the beehives during field studies. The infection rate ofN. ceranae increased proportionally with increasing temperature and humidity factors. Humidity was more effective than temperature on the infection rate ofN. ceranae. The seasonal activity ofN. ceranae was studied. The highest infection rates were observed in June and July. N. ceranae infection rate was higher in localities that were in low-altitude than in localities that were in high-altitude.

Which Endpoints Can Be Reliably Assessed in Semi-field Pollinator Species Testing without Estimating False Positive or False Negative？ MDD’s and Replicates Issue

Statistical power, number of replicates and experiment complexity of semi-field and field studies on Apis and non-Apis bee species has become a major issue after publication of the draft European Food Safety Authority （EFSA） Guidance on risk assessment of plant protection products （PPP） on bees （Apis mellifera, Bombus spp. and solitary bees）. According to this guidance document, field studies have to be designed to be able to detect significance differences as low as 7% for certain endpoints such as reduction in colony size. This will require an immense number of replicates which is obviously not feasible. In the present study, key endpoints such as mortality, termination rate and number of brood cells in honeybee studies, cocoon production and flight activity in solitary bee studies and number of gynes in bumble bee studies （just to mention some of the endpoints considered） in semi-field studies were analyzed, with Apis mellifera, Bombus terrestris and Osmia bicornis during the past five years （2013-2017）. The results indicate huge differences in the percentage minimal detectable differences （%MDDs） （MDD expressed as median of control value of the endpoint in percent） depending on endpoint and species tested. For honeybee semi-field studies, the lowest %MDDs recorded were between 10% and 15% for the endpoints foraging, number of brood cells and colony strength. The highest %MDDs were observed for the endpoint termination rate, with a %MDD of almost 50%. For the endpoints in bumble bee semi-field studies the %MDDs varied between 17% for bumble bee colony weight and 53% for average mortality during the exposure period in the tunnel. The %MDD for the number of gynes （young queens） was slightly below 25%. For the semi-field solitary bee test system, the %MDDs for the measured endpoints seem to be lower than those for the other two species tested. The %MDDs for the endpoints hatching of offspring, nest occupation and number of cocoons were 8%, 13% and 14%, respectively. Most of the %MDDs were between 10% and 30% indicating clearly that the currently performed experimental design for the semi-field pollinator studies allowed to determine relatively small effects on key study endpoints. The analysis indicated that for all the three bee species tested, the semi-field test design detected low %MDDs for most of the endpoints. It was also observed that detectable differences between the control and PPP treatments were much lower in semi-field test designs than in field studies with these bee species. The “perfect sample size” really does not exist but test design and statistical analysis can be adapted to lower the %MDDs. Measured and simulated （according to Student’s t-test-distribution） data and results showed that statistical evaluations parameter selection （e.g., alpha value）, data transformation （log10） and the number of replicates had a direct effect on the ability of the test design to detect lower or higher %MDD values. It could show that a change of alpha value from 0.05 to 0.1, increases the ability of the studies to detect lower %MDDs. For most of the measured endpoints, increasing the number of replicates e.g., from four to eight, improved the power of the test design by decreasing the %MDD. The reduction magnitude of the %MDD is dependent on the endpoint and selection of statistical parameters such as the alpha value. Parameters that display effects at a biologically relevant scale will be a better indicator for effects than parameters that are able to detect minor differences that are not biologically relevant.

A Survey of Pesticide Residues in Pollen Collected by Honey Bees from Four Different Types of Agricultural Production Areas in Slovenia

150 pollen samples collected between the years 2009 and 2011 from 30 locations, divided according to agricultural practice were analysed for the presence of pesticides. In 2009, residues were found in pollen samples from twelve locations （40%）, in 2010, in samples from four locations （13.3%） and in 2011, in samples from three locations （10%）. Altogether 15 different residues in pollen were found in 2009, two in year 2010 and five in 2011. Residues found in the pollen samples ranged from 0.001 mg/kg to 76 mg/kg. The highest number （twelve） of residues was found in pollen from the intensive fruit farming locations and from the intensive viticulture locations （eight）. The residues were mostly from the fungicide group （69%）. Insecticide residues （chlorpyrifos-ethyl, methoxyfenozide, thiacloprid） were found in the pollen samples from eight locations. Residues in the pollen did not statistically influence the development of the honeybee colonies or infestation rate of Nosema （Nosema spp.）, or viruses ABPV （acute bee paralysis virus）, SBV （sacbrood virus）, DWV （deformed wing virus）, and BQCV （black queen cell virus））.