Acessibilidade / Reportar erro

Using Citronella to Protect Bees (honeybee Apis mellifera L.) from certain Insecticides and Their Nano Formulations

Uso da citronela para proteger as abelhas (Apis mellifera L.) de certos inseticidas e suas nanoformulações

Abstract

Experiments were performed investigating citronella (Cymbopogon winterianus Jowitt) as a repellent to honeybee Apis mellifera (L.) (Hymenoptera: Apidae) in Egypt, it was conducted in laboratory in the Department of Entomology and Pesticides Science, Faculty of Agriculture, Cairo University, to check long-term survival of honeybee when exposed to different nano insecticides alone or combined with citronella at the same examination box for each. In this study, we used a modeling approach regarding survival data of caged worker bees under chronic exposure to four insecticides (Chloropyrophos, Nano-chloropyrophos Imidacloprid, Nano-Imidacloprid) each of them was supplemented in a box alone and in combination with citronella. Having three replicates and five concentrations (100, 200, 300, 400 and 500 ppm). Laboratory bioassay of these insecticides showed that chloropyrophos and nano chloropyrophos were the most toxic at their high dose (500 ppm) with LT50 of 120.98 and 122.02 followed by 132.14 and 136.5 minutes for Imidacloprid and Nano-Imidacloprid, respectively. No consumption occurred by bees to mixed sugar syrup with insecticides in all treatments when citronella was added. These data highly recommended that adding citronella is very effective when nicotinoid pesticides are used to longevity honeybee life and keep bee safe.

Keywords:
honeybee; Nano-Chlorpyrifos; Nano-Imidacloprid; citronela; toxicity

Resumo

Foram realizados experimentos para investigar a citronela (Cymbopogon winterianus Jowitt) como repelente de abelhas Apis mellifera (L.) (Hymenoptera: Apidae) no Egito, conduzidos no laboratório do Departamento de Entomologia e Ciência de Pesticidas, da Faculdade de Agricultura, da Universidade do Cairo, e verificar a sobrevivência a longo prazo das abelhas quando expostas a diferentes nanoinseticidas isoladamente ou combinados com citronela na mesma caixa de exame para cada um. Neste estudo, usamos uma abordagem de modelagem em relação aos dados de sobrevivência de abelhas operárias enjauladas sob exposição crônica a quatro inseticidas (clorpirifós, nanoclorpirifós, imidacloprida e nanoimidacloprida), e cada um deles foi suplementado em uma caixa e em combinação com citronela, tendo três repetições e cinco concentrações (100, 200, 300, 400 e 500 ppm). O bioensaio em laboratório desses inseticidas mostrou que clorpirifós e nanoclorpirifós foram os mais tóxicos em altas doses (500 ppm) com LT50 de 120,98 e 122,02, seguidos por 132,14 e 136,5 minutos para imidacloprida e nanoimidacloprida, respectivamente. Não houve consumo pelas abelhas do xarope de açúcar misto com inseticidas em todos os tratamentos quando a citronela foi adicionada. Esses dados recomendam a adição de citronela, sendo muito eficaz quando pesticidas nicotinoides são utilizados para longevidade das abelhas e para mantê-las seguras.

Palavras-chave:
abelha; Nanoclorpirifós; Nanoimidacloprida; citronela; toxicidade

1. Introduction

A lot of reports suggesting that exposure to citronella odor repel honeybees (Malerbo-Souza and Nogueira-Couto 2004MALERBO-SOUZA, D.T. and NOGUEIRA-COUTO, R.H., 2004. Efficiency of n-Octyl-Acetate, 2-Heptanone andcitronellal in repelling bees from basil (Ocimum sellowii – Labiatae). Brazilian Archives of Biology and Technology, vol. 47, no. 1, pp. 121-125. http://dx.doi.org/10.1590/S1516-89132004000100016.
http://dx.doi.org/10.1590/S1516-89132004...
; Abramson et al., 1999aABRAMSON, C.I., AQUINO, I.S., RAMALHO, F. and PRICE, J., 1999a. The effect of insecticides on learning in the Africanized honey bee (Apis mellifera L.). Archives of Environmental Contamination and Toxicology, vol. 37, no. 4, pp. 529-535. http://dx.doi.org/10.1007/s002449900548. PMid:10508901.
http://dx.doi.org/10.1007/s002449900548...
, Abramson and Aquino 2002aABRAMSON, C.I. and AQUINO, I.S., 2002a. Behavioral studies of learning in the Africanized honey bee (Apis mellifera L.). Brain Behaviour Evolution Journal., vol. 59, no. 1-2, pp. 68-86. http://dx.doi.org/10.1159/000063734. PMid:12097861.
http://dx.doi.org/10.1159/000063734...
, bABRAMSON, C.I. and AQUINO, I.S., 2002b. A scanning electron microscope atlas of the Africanized honey bee (Apis mellifera L.): Photographs for the general public. Campina Grande, Arte Express, 155 p.; Aquino et al., 2004AQUINO, I.S., ABRAMSON, C.I., SOARES, A.E., FERNANDES, A.C. and BENBASSAT, D., 2004. Classical conditioning of proboscis extension in harnessed Africanized honey bee queens (Apis mellifera L.). Psychological Reports Journal., vol. 94, no. 3 Pt 2, pp. 1221-1231. http://dx.doi.org/10.2466/pr0.94.3c.1221-1231. PMid:15362396.
http://dx.doi.org/10.2466/pr0.94.3c.1221...
). In addition to helping ensure a diverse supply of food for humans, pollination plays a critical role in providing the basis for essential ecosystem productivity and services (Kevan et al., 1990KEVAN, P.G., CLARK, E.A. and THOMAS, V.G., 1990. Pollination: A crucial ecological and mutualistic link inagro-forestry and sustainable agriculture. Proc Entomology Society Ont, vol. 121, pp. 43-48.; Kevan, 1999KEVAN, P.G., 1999. Pollinators as bioindicators of the state of the environment; species, activity anddiversity. Agriculture Ecosystem Environmental Journal., vol. 74, no. 1-3, pp. 373-393. http://dx.doi.org/10.1016/S0167-8809(99)00044-4.
http://dx.doi.org/10.1016/S0167-8809(99)...
., Antonini et al., 2006ANTONINI, Y., COSTA, R.G. and MARTINS, R.P., 2006. Floral preferences of a neotropical stingless bee, Melipona quadrifasciata Lepeletier (Apidae: Meliponina) in an urban forest fragment. Brazilian Journal of Biology = Revista Brasileira de Biologia, vol. 66, no. 2A, pp. 463-471. http://dx.doi.org/10.1590/S1519-69842006000300012. PMid:16862301.
http://dx.doi.org/10.1590/S1519-69842006...
). There is concern about potential adverse effects of pesticides on pollinators (EFSA, 2012EFSA, 2012. EFSA panel on plant protection products and their residues (PPR); scientific opinion on the science behind the development of a risk assessment of plant protection products on bees (Apis mellifera, Bombus spp. and solitary bees). EFSA Journal, vol. 10, no. 5, pp. 2668. http://dx.doi.org/10.2903/j.efsa.2012.2668.
http://dx.doi.org/10.2903/j.efsa.2012.26...
; National Research Council, 2007NATIONAL RESEARCH COUNCIL, 2007. Status of pollinators in North America. Washington, DC: The National Academies Press, 322 pp). Considerable effort has been directed at finding a honeybee repellent because of public safety issues (Abramson et al. 1997aABRAMSON, C.I., AQUINO, I.S., AZEREDO, G.A., FILHO, J.R.M. and PRICE, J.M., 1997a. The attraction of Africanized honey bees (Apis mellifera L.) to soft drinks and perfumes. The Journal of Genetic Psychology, vol. 124, no. 2, pp. 166-181. http://dx.doi.org/10.1080/00221309709595515.
http://dx.doi.org/10.1080/00221309709595...
), the possibility of providing researchers interested in reducing the effects of harmful agrochemicals (Atkins Jr. et al., 1975a, 1975b; Mayer, 1997MAYER, D.F., 1997. Effects of methyl salicylate on honey bee (Apis mellifera L.) foraging. New Zealand Journal of Crop and Horticultural Science, vol. 25, no. 3, pp. 291-294. http://dx.doi.org/10.1080/01140671.1997.9514018.
http://dx.doi.org/10.1080/01140671.1997....
). There are several studies in the literature suggesting other repellent to honeybees (Blum et al., 1978BLUM, M.S., FALES, H.M., TUCKER, K.W. and CPLLINS, A.M., 1978. Chemistry of the sting apparatus of theworker honeybee. Journal of Apicultural Research, vol. 17, no. 4, pp. 218-221. http://dx.doi.org/10.1080/00218839.1978.11099929.
http://dx.doi.org/10.1080/00218839.1978....
; Free, 1987FREE, J.B., 1987. Pheromones of social bees. New York: Chapman & Hall, 236 p.; Free et al., 1989FREE, J.B., FERGUSON, A.W. and SIMPKINS, J.R., 1989. Honeybee responses to chemical components from the worker sting apparatus mandibular glands in field tests. Journal of Apicultural Research, vol. 28, no. 1, pp. 7-21. http://dx.doi.org/10.1080/00218839.1989.11100814.
http://dx.doi.org/10.1080/00218839.1989....
). Impact of pesticides as a bioassay to screen for adulterated honey was assessed (Abramson et al.; 1999bABRAMSON, C.I., AQUINO, I.S., RAMALHO, F.S. and PRICE, J.M., 1999b. The effect of insecticides on learningin the Africanized honey bee (Apis mellifera L.). Archives of Environmental Contamination and Toxicology, vol. 37, no. 4, pp. 529-535. http://dx.doi.org/10.1007/s002449900548. PMid:10508901.
http://dx.doi.org/10.1007/s002449900548...
; Abramson et al., 2006ABRAMSON, C.I., SINGELTON, J.B., WILSON, M.K., WANDERLAY, P.A., RAMALHO, F.S. and MICHALUK, L.M., 2006. The effect of an organic pesticide on mortality and learning in Africaned honey bees (Apis melliferaL.) in Brasil. American Journal of Environmental Sciences, vol. 2, pp. 37-44.). In this report, the ability of citronella odor to repel honeybee under laboratory conditions was tested. We believe this approach can serve as a model for testing repellents in honeybees. However, the number of managed honeybee colonies has declined in Europe and the United States (Dainat et al., 2011DAINAT, B., VANENGELSDORP, D. and NEUMANN, P., 2011. Colony collapse disorder in Europe. Environmental Microbiology Reports, vol. 4, no. 1, pp. 123-125. http://dx.doi.org/10.1111/j.1758-2229.2011.00312.x. PMid:23757238.
http://dx.doi.org/10.1111/j.1758-2229.20...
; vanEngelsdorp et al., 2011VANENGELSDORP, D., HAYES, J., UNDERWOOD, R., CARON, D. and PETTIES, J., 2011. A survey of managed honey bee colony losses in the USA, Fall 2009 to Winter 2010. Journal of Apicultural Research, vol. 50, no. 1, pp. 1-10. http://dx.doi.org/10.3896/IBRA.1.50.1.01.
http://dx.doi.org/10.3896/IBRA.1.50.1.01...
) due to multiple factors (Dainat et al., 2012DAINAT, B., EVANS, J.D., CHEN, Y.P., GAUTHIER, L. and NEUMANN, P., 2012. Predictive markers of honey bee colony collapse. PLoS One, vol. 7, no. 2, pp. e32151. http://dx.doi.org/10.1371/journal.pone.0032151. PMid:22384162.
http://dx.doi.org/10.1371/journal.pone.0...
; van Lexmond et al., 2015VAN LEXMOND, M.B., BONMATIN, J.M., GOULSON, D. and NOOME, D.A., 2015. Worldwide integrated assessment on systemic pesticides: Global collapse of the entomofauna: Exploring the role of systemic insecticides. Environmental Science and Pollution Research, vol. 22, pp. 1-4.). Recently, attention has focused on neonicotinoid pesticides like imidacloprid, which can impair individual and colony fitness at even sublethal doses (Desneux et al., 2007DESNEUX, N., DECOURTYE, A. and DELPUECH, J.M., 2007. The sublethal effects of pesticides on beneficial arthropods. Annual Review of Entomology, vol. 52, no. 1, pp. 81-106. http://dx.doi.org/10.1146/annurev.ento.52.110405.091440. PMid:16842032.
http://dx.doi.org/10.1146/annurev.ento.5...
; Goulson, 2013GOULSON, D., 2013. An overview of the environmental risks posed by neonicotinoid insecticides. Journal of Applied Ecology, vol. 50, no. 4, pp. 977-987. http://dx.doi.org/10.1111/1365-2664.12111.
http://dx.doi.org/10.1111/1365-2664.1211...
; Henry et al., 2012HENRY, M., BEGUIN, M., REQUIER, F., ROLLIN, O., ODOUX, J.F., AUPINEL, P., APATEL, J., TCHAMITCHIAN, S. and DECOURTYE, A., 2012. A common pesticide decreases foraging success and survival in honey bees. Science Journal, vol. 336, no. 6079, pp. 348-350. http://dx.doi.org/10.1126/science.1215039.
http://dx.doi.org/10.1126/science.121503...
; Sanchez-Bayo, 2011SÁNCHEZ-BAYO, S., 2011. Insecticides Mode of Action in Relation to Their Toxicity to Non-Target Organisms. J Environment Analytic Toxicol, vol. S4. http://dx.doi.org/10.4172/2161-0525.S4-002
https://doi.org/10.4172/2161-0525.S4-002...
). Imidacloprid is a systemic insecticide that spreads throughout all plant tissues and is found in nectar and pollen at concentrations up to 50 ppm (Goulson, 2013GOULSON, D., 2013. An overview of the environmental risks posed by neonicotinoid insecticides. Journal of Applied Ecology, vol. 50, no. 4, pp. 977-987. http://dx.doi.org/10.1111/1365-2664.12111.
http://dx.doi.org/10.1111/1365-2664.1211...
). Imidacloprid can also be found in water with 8% (Johnson and Pettis, 2014JOHNSON, J.D. and PETTIS, J.S., 2014. A survey of imidacloprid levels in water sources potentially frequented by honeybees (Apis mellifera) in the eastern USA. Water Air Soil Pollitten., vol. 225, no. 11, pp. 2127. http://dx.doi.org/10.1007/s11270-014-2127-2. PMid:25477698.
http://dx.doi.org/10.1007/s11270-014-212...
). Chlorpyrifos is an organophosphorus insecticide control a wide variety of foliage- and soil-borne insect pests on a variety of food and feed crops (Solomon et al., 2014SOLOMON, K.R., WILLIAMS, W.M., MACKAY, D., PURDY, J., GIDDINGS, J.M. and GIESY, J.P., 2014. Properties and uses of chlorpyrifos in the United States. Reviews of Environmental Contamination and Toxicology, vol. 231, pp. 13-34. http://dx.doi.org/10.1007/978-3-319-03865-0_2. PMid:24723132.
http://dx.doi.org/10.1007/978-3-319-0386...
). Rising interest occurred recently with nanopesticides. Nano-imidacloprid has been used to control several insects (Sabbour, 2015SABBOUR, M.M., 2015. Efficacy of some nano-Imidacloprid against red flour beetle Tribolium confusum (Coleoptera Tenebrionidae) under laboratory and stored conditions. Advances in Biochemistry & Biotechnology. vol. 1, pp. 1-13.; Assemi et al., 2014ASSEMI, H., SAJJADIE, A. and NAGHIZADEH, F., 2014. Investigation of different values of nano imidacloprid for control Tabaco aphid Myzus persicae nicotianae in Laboratory. Agrotechnology Journal., vol. 3, pp. 128. http://dx.doi.org/10.4172/2168-9881.1000128.
http://dx.doi.org/10.4172/2168-9881.1000...
). This study refers to using citronella could be very effective especially when we use nicotinoides pesticides, in order to repel bee and keep them safe.

2. Material and Methods

2.1. Insect

Honeybee Apis mellifera workers obtained from apiary at Faculty of agriculture, Cairo University.

Preparation of Nano Pesticides

Nanochlorpyrifos and nanoimidacloprid were both prepared at Nano Tech Company, Dreamland, 6th October City, Egypt according to the method described by Guan et al. (2008)GUAN, H., CHI, D., YU, J. and LI, X., 2008. A novel photodegradable insecticide: Preparation, characterization and properties evaluation of nano-Imidacloprid. Pesticide Biochemistry and Physiology, vol. 92, no. 2, pp. 83-91. http://dx.doi.org/10.1016/j.pestbp.2008.06.008.
http://dx.doi.org/10.1016/j.pestbp.2008....
. The Fourier Transform Technique (for Advanced Readers) Fourier Transform type (Mac Company) was used; the technique was according to (Griffith and Fuller, 1982GRIFFITH, P.R. and FULLER, M.P., 1982. Advances in Infrared and Raman Spectroscopy. Clark, R.J.H. and Hester, R. E., eds, Heydon & Sons: London, vol. 9, pp. 63-129.).

2.2. TEM Analysis

The shape and size of both nano chlorpyrifos and nano imidacloprid were analysed and observed under Transmission Electron Microscope (TEM), JEOL model 3010 Philips CM-200, Japan operated at 120 kv, they were prepared according to Parveen et al. (2014)PARVEEN, S.A., MARTIN, R.J. and SAHAYARAJ, K., 2014. Terminals Chebula Retz gallic acid – Based silver nanoparticles and their antiphytopathogenic activity. Journal of Biopesticides, vol. 7, pp. 1-6..

2.3. Bioassay

Toxicity was studied with special wooden boxes of diameters (75 cm width and 80 cm height surrounded with metal net from all sides). In this method, we supplied each cage with two petri dishes 10 cm diameter, each one contained 5ml of each insecticide or insecticide with citronella with recommended repel concentration. Five honeybees’ workers were introduced, along with one control box free of any pesticides but it containing distilled water and sugar (1:1). This treatment was replicated three times.

Also, mortality percentage was determined after 30, 60, 90, 120 seconds and so on. Honeybee workers were kept in cage for very shorter periods of time (up to 24 hours). Methodology for assay was developed in the lab, so there is no reference in this regard. The boxes were placed at temperature 25 ± 10 ºC and 50 ± 5% humidity. Food consummation was determined through measurements of syrup after 2 hours from each petri dish. Besides our team designed direction box attitude (Figure 1) where we join tow boxes with cylindrical bridge with open passage (path) from the top to release bees, one of the boxes contained Nano-Imidacloprid 500 ppm and the other one contained the same insecticide with citronella and bees released in the middle to the bridge and let to choose any direction freely where to go.

Figure 1
Cage to investigate bee food preferences where A contains pesticide alone and B contains Pesticide and citronella mixed together and C a passage or path where bee let go free to A or B

2.4. Statistical analysis

LD50 values were estimated by plotting log dose versus probit plus five mortality (Sokal and Rohlf, 1995SOKAL, R.R. and ROHLF, F.J., 1995. Biometry The Principles and Practice of Statistics in Biological Research. 3rd ed. New York: W.H. Freeman and Co.; Finney, 1971FINNEY, D., 1971. Probit analysis. Cambridge, UK: Cambridge University Press, 256 p.; Microsoft Excel, 1997MICROSOFT EXCEL, 1997. Microsoft Excel. Redmond, WA: Microsoft.).

3. Results and Discussion

The sudden disappearance of honeybee from hives has been called “Disappearing Disease” (Wilson and Menapace, 1979WILSON, W.T. and MENAPACE, D.M., 1979. Disappearing disease of honey bees: A survey of the United States. American Bee Journal, vol. 119, pp. 184-186.). It was noted and referred to as Colony Collapse Disorder (CCD) by researcher, also corresponded to increased use of neonicotinoid pesticides (Johnson et al., 2010JOHNSON, R.M., ELLIS, M.D., MULLIN, C.A. and FRAZIER, M., 2010. Pesticides and honey bee toxicity–USA. Apidologie, vol. 41, no. 3, pp. 312-331. http://dx.doi.org/10.1051/apido/2010018.
http://dx.doi.org/10.1051/apido/2010018...
; Cresswell et al., 2012CRESSWELL, J.E., DESNEUX, N. and VANENGELSDORP, D., 2012. Dietary traces of neonicotinoid pesticides as a cause of population declines in honey bees: An evaluation by Hill’s epidemiological criteria. Pest Management Science Journal, vol. 68, no. 6, pp. 819-827. http://dx.doi.org/10.1002/ps.3290. PMid:22488890.
http://dx.doi.org/10.1002/ps.3290...
). This has led to speculation that there is a causative relationship between increasing using by neonicotinoids and widely decline in bee populations (Suryanarayanan, 2013SURYANARAYANAN, S., 2013. Balancing control and complexity in field studies of neonicotinoids and honey bee health. Insects, vol. 4, no. 1, pp. 153-167. http://dx.doi.org/10.3390/insects4010153. PMid:26466800.
http://dx.doi.org/10.3390/insects4010153...
). However, it is important to look at all the variables associated with CCD. Malerbo-Souza and Nogueira-Couto (2004)MALERBO-SOUZA, D.T. and NOGUEIRA-COUTO, R.H., 2004. Efficiency of n-Octyl-Acetate, 2-Heptanone andcitronellal in repelling bees from basil (Ocimum sellowii – Labiatae). Brazilian Archives of Biology and Technology, vol. 47, no. 1, pp. 121-125. http://dx.doi.org/10.1590/S1516-89132004000100016.
http://dx.doi.org/10.1590/S1516-89132004...
found that spraying citronella temporary decrease in the number of bees visiting a test site (Harpaz and Lensky, 1959HARPAZ, I. and LENSKY, Y., 1959. Experiments on bee repellents. Bee World, vol. 40, no. 6, pp. 146-153. http://dx.doi.org/10.1080/0005772X.1959.11096718.
http://dx.doi.org/10.1080/0005772X.1959....
). It has been known for some time that honeybee learn to associate a floral scent with (Menzel et al., 1993MENZEL, R., GREGGERS, U. and HAMMER, M., 1993. Functional organization of appetitive learning andmemory in a generalist pollinator, the honey bee. In: D.R. PAPAJ and A.C. LEWIS, eds. Insectlearning: Ecological and evolutionary perspectives. New York: Chapman & Hall, pp. 79-125.). It is entirely likely that the odor of the test repellent is the reason of this. (Abramson et al., 1996ABRAMSON, C.I., BUCKBEE, D.A., EDWARDS, S. and BOWE, K., 1996. A demonstration of virtual reality in freeflying honey bees: Apis mellifera. Physiological Behavior Journal., vol. 59, no. 1, pp. 39-43. http://dx.doi.org/10.1016/0031-9384(95)02023-3. PMid:8848488.
http://dx.doi.org/10.1016/0031-9384(95)0...
, Silva and Rebêlo, 2002SILVA, F. S. and REBÊLO, J. M. M., 2002. Population dynamics of euglossinae bees(Hymenoptera, Apidae) in an earlysecond-growth forest of cajual island,in the state of Maranhão, Brazil. Brazilian Journal of Biology, vol. 62, 1, pp. 15-23.). Our belief on ongoing research is increasing our understanding of the impact of these types of pesticides on bees. For now, the best recommendation is to carefully follow the product label, be judicious in your application, and avoid applying any insecticide product, if necessary you should it is useful to use citronella as a repellent so you don’t affect the bees.

3.1. Food consumption and wheight

All treatments were detected for syrup consumption and preference of bee to eat, and it was found that all treatments concluded citronella was very detritus, where it scored zero consumption in comparison to other treatments where bee preferred to feed in control treatments followed by Imidacloprid, Chloropyrophos, Nano-Imidacloprid and Nano-Chloropyrophos; respectively, also in direction cage experiment 97.3% of total released bees moved towards the box containing only pesticide avoiding the other box that contain pesticide mixed with citronella.

3.2. Legs Shrinkage

The importance of examine this parameter; attributed to that Imidacloprid is considered neurotoxin where it affects mostly the synapsis in CNS of honeybee. Results of Nano-Imidacloprid showed that highly affected on worker legs where it recorded 100.00, 90.00, 100.00, 89.47 and 91.66% with nano Imidacloprid 100, 200, 300, 400 and 500 ppm, respectively. Where it gave 63.63% with Imidacloprid and 100.00 with Chloropyrifos. These ratio decreases significantly where Nano-chloropyrophos was examined, it gave 0.00, 75.00, 63.15, 58.33 and 100.00% with 100, 200, 300, 400 and 500 ppm concentrations, respectively. On the other hand, for sugar syrup that formulated from distilled water, data obtained showed that shrinkage in legs was limited where it gave 11.11% only, (Figure 2)

Figure 2
Leg shortage in honey bee workers caused by (A) Imidacloprid and nano Imidacloprid and (B) Chloropyrophos and nano chloropyrophos.

3.3. Circulation Degree of the Body

Nano-Imidacloprid results showed that highly affected on body balling of honeybee workers where it recorded 88.00, 100.00, 100.0, 90.00 and 91.00% with Nano-Imidacloprid 100, 200, 300, 400 and 500 ppm, respectively. Where it gave 100% with both Imidacloprid and Chloropyrifos, this ratio decreased significantly where Nano- Chloropyrophos was examined; it gave 10.00, 62.50, 57.14, 78.94 and 83.33% with 100, 200, 300, 400 and 500 ppm concentrations, respectively. On the other hand, for syrup formulated from distilled water and sugar there were no shortage (balling) occurred at all worker members (Figure 3).

Figure 3
Body circulation in honey bee workers caused by (A) Imidacloprid and nano Imidacloprid and (B) Chloropyrophos and nano chloropyrophos.

3.4. Abdomen shortage

Imidacloprid can have delayed toxic effects in bees, particularly in chronic exposure (Rondeau et al., 2014RONDEAU, G., SÁNCHEZ-BAYO, F., TENNEKES, H.A., DECOURTYE, A., RAMÍREZ-ROMERO, R. and DESNEUX, N., 2014. Delayed and time-cumulative toxicity of imidacloprid in bees, ants and termites. Scientific Reports, vol. 4, pp. 1-8. PMid:24993452.). Their degradation was toxic to bees (Goulson, 2013GOULSON, D., 2013. An overview of the environmental risks posed by neonicotinoid insecticides. Journal of Applied Ecology, vol. 50, no. 4, pp. 977-987. http://dx.doi.org/10.1111/1365-2664.12111.
http://dx.doi.org/10.1111/1365-2664.1211...
). Imidacloprid is insecticidal because it is an agonist of insect nicotinic acetylcholine receptors (nAChRs) (Gauthier, 2010GAUTHIER, M. 2010. State of the art on insect nicotinic acetylcholine receptor function in learning and memory. In: S.H. THANY. Insect acetylcholine receptors. New York: Springer, pp. 97-115. http://dx.doi.org/10.1007/978-1-4419-6445-8_9.
http://dx.doi.org/10.1007/978-1-4419-644...
), which are found in honeybee (Jones et al., 2005JONES, J.C., HELLIWELL, P., BEEKMAN, M., MALESZKA, R. and OLDROYD, B.P., 2005. The effects of rearing temperature on developmental stability and learning and memory in the honey bee, Apis mellifera. Journal of Comparative Physiology, vol. 191, no. 12, pp. 1121-1129. http://dx.doi.org/10.1007/s00359-005-0035-z. PMid:16049697.
http://dx.doi.org/10.1007/s00359-005-003...
) and play a role in bee learning (Dacher and Gauthier, 2008DACHER, M. and GAUTHIER, M., 2008. Involvement of NO-synthase and nicotinic receptors in learning in the honey bee. Physiological behavior Journal. 95(1-2), 200-207.). Honeybee nAChRs play an important role in cholinergic neural signaling, and bees fed sublethal doses of imidacloprid down regulate nAChRs in their brains (Zhou et al., 2014ZHOU, T., SONG, H.L., WANG, Q., DAI, P.L., WU, Y.Y. and SUN, J.H., 2014. Effects of imidacloprid on the distribution of nicotine acetylcholine receptors in the brain of adult honeybee (Apis mellifera ligustica). Acta Entomology Sinica, vol. 56, pp. 1258-1266.). Imidacloprid has a wide variety of effects: brain cell death (Wu et al., 2014WU, Y.Y., ZHOU, T., WUBIE, A.J., WANG, Q., DAI, P.L. and JIA, H.R., 2014. Apoptosis in the nerve cells of adult honeybee (Apis mellifera ligustica) brain induced by imidacloprid. Acta Entomology Sinica, vol. 57, pp. 194-203.), impaired foraging (Schneider et al., 2012SCHNEIDER, C.W., TAUTZ, J., GRÜNEWALD, B. and FUCHS, S., 2012. RFID tracking of sublethal effects of two neonicotinoid insecticides on the foraging behavior of Apis mellifera. PLoS One, vol. 7, no. 1, pp. e30023. http://dx.doi.org/10.1371/journal.pone.0030023. PMid:22253863.
http://dx.doi.org/10.1371/journal.pone.0...
, decreased food uptake (Ramirez-Romero et al., 2005RAMIREZ-ROMERO, R., CHAUFAUX, J. and PHAM-DELEGUE, M.H., 2005. Effects of Cry1Ab protoxin, deltamethrin and imidacloprid on the foraging activity and the learning performances of the honeybee Apis mellifera, a comparative approach. Apidologie, vol. 36, no. 4, pp. 601-611. http://dx.doi.org/10.1051/apido:2005039.
http://dx.doi.org/10.1051/apido:2005039...
), diminished hive entrance activity (Decourtye et al., 2004aDECOURTYE, A., ARMENGAUD, C., RENOU, M., DEVILLERS, J., CLUZEAU, S., GAUTHIER, M. and PHAM-DELEGUE, M.H., 2004a. Imidacloprid impairs memory and brain metabolism in the honeybee (Apis mellifera L.). Pesticide Biochemistry and Physiology, vol. 78, no. 2, pp. 83-92. http://dx.doi.org/10.1016/j.pestbp.2003.10.001.
http://dx.doi.org/10.1016/j.pestbp.2003....
), reduced motor function (Lambin et al., 2001LAMBIN, M., ARMENGAUD, C., RAYMOND, S. and GAUTHIER, M., 2001. Imidacloprid‐induced facilitation of the proboscis extension reflex habituation in the honeybee. Archives of Insect Biochemistry and Physiology, vol. 48, no. 3, pp. 129-13. http://dx.doi.org/10.1002/arch.1065. PMid:11673842.
http://dx.doi.org/10.1002/arch.1065...
; Williamson and Wright, 2013WILLIAMSON, S.M. and WRIGHT, G.A., 2013. Exposure to multiple cholinergic pesticides impairs olfactory learning and memory in honeybees. The Journal of Experimental Biology, vol. 216, no. Pt 10, pp. 1799-1807. http://dx.doi.org/10.1242/jeb.083931. PMid:23393272.
http://dx.doi.org/10.1242/jeb.083931...
), impaired visual learning (Han et al., 2010HAN, P., NIU, C. Y., LEI, C. L., CUI, J. J. and DESNEUX, N., 2010. Use of an innovative T-tube maze assay and the proboscis extension response assay to assess sublethal effects of GM products and pesticides on learning capacity of the honey bee Apis mellifera L. Ecotoxin Journal, vol. 19, pp. 1612-1619.), decreased predator avoidance (Araújo et al., 2004ARAÚJO, E.D., COSTA, M., CHAUD-NETTO, J. and FOWLER, H.G., 2004. Body size and flight distance in stingless bees (Hymenoptera:meliponini): inference of flightrange and possible ecological implications. Brazilian Journal of Biology = Revista Brasileira de Biologia, vol. 64, no. 3B, pp. 563-568. http://dx.doi.org/10.1590/S1519-69842004000400003. PMid:15619994.
http://dx.doi.org/10.1590/S1519-69842004...
), impaired navigation to the nest (Fischer et al., 2014FISCHER, J., MÜLLER, T., SPATZ, A.K., GREGGERS, U., GRÜNEWALD, B. and MENZEL, R., 2014. Neonicotinoids interfere with specific components of navigation in honeybees. PLoS One, vol. 9, no. 3, pp. e91364. http://dx.doi.org/10.1371/journal.pone.0091364. PMid:24646521.
http://dx.doi.org/10.1371/journal.pone.0...
) flowers (Dukas and Morse, 2003DUKAS, R. and MORSE, D.H., 2003. Crab spiders affect flower visitation by bees. Oikos, vol. 101, no. 1, pp. 157-163. http://dx.doi.org/10.1034/j.1600-0706.2003.12143.x.
http://dx.doi.org/10.1034/j.1600-0706.20...
; Morse and Nowogrodzki, 1990MORSE, R.A. and NOWOGRODZKI, R., 1990. Honey bee pests, predators, and diseases. Ithaca, New York: Comstock Publishing Associates.; Reader et al., 2006READER, T., HIGGINSON, A.D., BARNARD, C.J. and GILBERT, F.S., 2006. The effects of predation risk from crab spiders on bee foraging behavior. Behavior Ecology Journal., vol. 17, no. 6, pp. 933-939. http://dx.doi.org/10.1093/beheco/arl027.
http://dx.doi.org/10.1093/beheco/arl027...
). This is a complementary parameter where it gave obvious picture on situation of muscles, which really were affected, that in all treatments no significant effects were found in abdomen shortage (Figure 4).

Figure 4
Abdomen shortage in honey bee workers caused by (A) Imidacloprid and nano Imidacloprid and (B) Chloropyrophos and nano chloropyrophos.

3.5. Wing horizontal

The normal attitude to have 0.00% wing horizontal manner. As it is illustrated in Figure 5 where this parameter gave 0.00% in case of syrup free of pesticide, where it gave 63.63% with Imidacloprid and 33.33, 20.00, 12.5, 25.0, and 9.09% with Nano- Imidacloprid concentrations (100, 200, 300, 400 and 500, respectively). In contrast, this ratio increased gradually with chloropyrophos where it gave 100% with chloropyrophos, and 0.00, 75.00, 14.28 and 63.15 and 58.33% with 100, 200, 300, 400 and 500 ppm of nano-chloropyrophos concentrations, respectively, (Figure 5).

Figure 5
Wing horizontal in honey bee workers caused by (A) Imidacloprid and nano Imidacloprid and (B) Chloropyrophos and nano chloropyrophos.

3.6. The effect on mouth parts

This parameter showed highly killing speed where bee dies through having syrup quickly, Figure 6 shows zero % with syrup free of any pesticides. Whereas it was greatly increased with the lowered concentrations of Nano-Imidacloprid, where it recorded 88.88, 80.00, 50.00, 65.00 and 66.66% with concentrations 100, 200, 300, 400 and 500 ppm, respectively. And 36.36% with Imidacloprid.

Figure 6
Mouthparts out and straight in honey bee workers caused by (A) Imidacloprid and nano Imidacloprid and (B) Chloropyrophos and nano chloropyrophos.

This effect decreased significantly with chloropyrophos which scored 18.18% only, and its nano form recorded 0.00, 75.00, 14.28, 63.15 and 58.33% with 100, 200, 300, 400 and 500 ppm, respectively.

3.7. The fourier transform technique

The nano-pesticides were examined for its activity stability according to (Griffith and Fuller, 1982GRIFFITH, P.R. and FULLER, M.P., 1982. Advances in Infrared and Raman Spectroscopy. Clark, R.J.H. and Hester, R. E., eds, Heydon & Sons: London, vol. 9, pp. 63-129.; Griffith and Haseth, 1986GRIFFITH, P.R. and HASETH, J.A., 1986. Fourier transform infrared spectrometry. New York: Wiley and Sons.; Calvert et al., 1993CALVERT, J.G., HEYWOOD, J.B., SAWYER, R.F. and SEINFELD, J.H., 1993. A chieving acceptable air quality: some reflections on controlling vehicle emissions. Science Journal., vol. 261, no. 5117, pp. 37-45. PMid:17750544.; Reuter et al., 1998REUTER, J.E., ALLEN, B.C., RICHARDS, R.C., PANKOW, J.F., GOLDMAN, C.R., SCHOLL, R.L. and SEYFRIED, J.S., 1998. Concentrations, souPerces, and fate of the gasoline oxygenate methyl/e/7-butyl ether (MTBE) in a multiple-use lake. Environmental Science & Technology, vol. 32, no. 23, pp. 3666-3672. http://dx.doi.org/10.1021/es9805223.
http://dx.doi.org/10.1021/es9805223...
; Johnson et al., 2000JOHNSON, R., PANKOW, J., BENDER, D., PRICE, C. and ZOGORISKI, J., 2000. MtbeT, to what extent will past releases contaminate community Water supply welly. Environmental Science Tecnology Journal, vol. 34, no. 9, pp. 210a-217a. http://dx.doi.org/10.1021/es003268z.
http://dx.doi.org/10.1021/es003268z...
). Data given in Figures 7 and 8 revealed that nano-chlorpyrifos almost the same peaks as compared to with chlorpyrifos while nano-imidacloprid peaks almost reduced to a half when compared to active peaks with imidacloprid.

Figure 7
FTIR absorption curves where X represent wave length and Y represent absorption in nanometre (7A, 7C, 7E and 7G) nano Imidacloprid and (7B, 7D, 7F and 7H) represent Imidacloprid
Figure 8
FTIR absorption curves where X represent wave length and Y represent absorption in nanometre (8A,8 C, 8E and8 G) nano Chloropyrophos and (8B,8 D, 8F and 8H) represent Chloropyrophos.

3.8. TEM analysis

Both of imidacloprid and chlorpyrifos nanoparticles revealed nano size where the mean of counted particles were less than 100nm for nano chlorpyrifos (with median range 44.5nm) and nano imidacloprid (with median range 88.7nm) (Figure 9), once nutrition; Respectively. Metabolites of imidacloprid were found in honeybee (Nauen et al., 1998NAUEN, R., HUNGENBERG, H., TOLLO, B., TIETJEN, K. and ELBERT, A., 1998. Antifeedant effect, biologica l efficacy and high affinity binding of imidacloprid to acetylcholine receptors in Myzus persicae and Myzus nicotianae. Pesticides Science Journal, vol. 53, no. 2, pp. 133-140. http://dx.doi.org/10.1002/(SICI)1096-9063(199806)53:2<133::AID-PS756>3.0.CO;2-D.
http://dx.doi.org/10.1002/(SICI)1096-906...
; Decourtye et al., 2004bDECOURTYE, A., DEVILLERS, J., CLUZEAUl, S., CHARRETON, M., and PHAM-DELEGUE M.H., 2004b. Effects of Imidacloprid and deltamethrin on associative learning in honeybees under semifield and laboratory conditions. Ecotoxicology and Environmental Safety Journal, vol. 57, pp. 410-419.). Yamamoto et al. (1998)YAMAMOTO, I., TOMIZAWA, M., SAITO, T., MIYAMOTO, T., WALCOTT, E.C. and SUMIKAWA, K., 1998. Structural factors contributing to insecticidal and selective actions of neonicotinoids. Archives of Insect Biochemistry and Physiological Journal, vol. 37, no. 1, pp. 24-32. http://dx.doi.org/10.1002/(SICI)1520-6327(1998)37:1<24::AID-ARCH4>3.0.CO;2-V. PMid:9397513.
http://dx.doi.org/10.1002/(SICI)1520-632...
findings suggest that metabolism and detoxification pathways may vary between insect species which can affect insect susceptibility to neonicotinoids. We provide the first evidence that the neonicotinoid, nano-Imidacloprid, chloropyrophos or nano-chloropyrophos can significantly kill honeybee, where nano chloropyrophos, chloropyrophos were less toxic generally in comparison to nano-Imidacloprid and Imidacloprid, and that adding citronella lead to complete avoid to the syrup. We are suggesting that adding citronella and in further investigations field evaluation should be applied and maybe other repellants should be examined.

Figure 9
TEM image showing synthesized A, Imidacloprid nano particles and B, chloropyrophos nano particles.
  • (With 9 figures)

References

  • ABRAMSON, C.I. and AQUINO, I.S., 2002a. Behavioral studies of learning in the Africanized honey bee (Apis mellifera L.). Brain Behaviour Evolution Journal., vol. 59, no. 1-2, pp. 68-86. http://dx.doi.org/10.1159/000063734 PMid:12097861.
    » http://dx.doi.org/10.1159/000063734
  • ABRAMSON, C.I. and AQUINO, I.S., 2002b. A scanning electron microscope atlas of the Africanized honey bee (Apis mellifera L.): Photographs for the general public Campina Grande, Arte Express, 155 p.
  • ABRAMSON, C.I., AQUINO, I.S., AZEREDO, G.A., FILHO, J.R.M. and PRICE, J.M., 1997a. The attraction of Africanized honey bees (Apis mellifera L.) to soft drinks and perfumes. The Journal of Genetic Psychology, vol. 124, no. 2, pp. 166-181. http://dx.doi.org/10.1080/00221309709595515
    » http://dx.doi.org/10.1080/00221309709595515
  • ABRAMSON, C.I., AQUINO, I.S., RAMALHO, F. and PRICE, J., 1999a. The effect of insecticides on learning in the Africanized honey bee (Apis mellifera L.). Archives of Environmental Contamination and Toxicology, vol. 37, no. 4, pp. 529-535. http://dx.doi.org/10.1007/s002449900548 PMid:10508901.
    » http://dx.doi.org/10.1007/s002449900548
  • ABRAMSON, C.I., AQUINO, I.S., RAMALHO, F.S. and PRICE, J.M., 1999b. The effect of insecticides on learningin the Africanized honey bee (Apis mellifera L.). Archives of Environmental Contamination and Toxicology, vol. 37, no. 4, pp. 529-535. http://dx.doi.org/10.1007/s002449900548 PMid:10508901.
    » http://dx.doi.org/10.1007/s002449900548
  • ABRAMSON, C.I., BUCKBEE, D.A., EDWARDS, S. and BOWE, K., 1996. A demonstration of virtual reality in freeflying honey bees: Apis mellifera. Physiological Behavior Journal., vol. 59, no. 1, pp. 39-43. http://dx.doi.org/10.1016/0031-9384(95)02023-3 PMid:8848488.
    » http://dx.doi.org/10.1016/0031-9384(95)02023-3
  • ABRAMSON, C.I., SINGELTON, J.B., WILSON, M.K., WANDERLAY, P.A., RAMALHO, F.S. and MICHALUK, L.M., 2006. The effect of an organic pesticide on mortality and learning in Africaned honey bees (Apis melliferaL.) in Brasil. American Journal of Environmental Sciences, vol. 2, pp. 37-44.
  • ANTONINI, Y., COSTA, R.G. and MARTINS, R.P., 2006. Floral preferences of a neotropical stingless bee, Melipona quadrifasciata Lepeletier (Apidae: Meliponina) in an urban forest fragment. Brazilian Journal of Biology = Revista Brasileira de Biologia, vol. 66, no. 2A, pp. 463-471. http://dx.doi.org/10.1590/S1519-69842006000300012 PMid:16862301.
    » http://dx.doi.org/10.1590/S1519-69842006000300012
  • AQUINO, I.S., ABRAMSON, C.I., SOARES, A.E., FERNANDES, A.C. and BENBASSAT, D., 2004. Classical conditioning of proboscis extension in harnessed Africanized honey bee queens (Apis mellifera L.). Psychological Reports Journal., vol. 94, no. 3 Pt 2, pp. 1221-1231. http://dx.doi.org/10.2466/pr0.94.3c.1221-1231 PMid:15362396.
    » http://dx.doi.org/10.2466/pr0.94.3c.1221-1231
  • ARAÚJO, E.D., COSTA, M., CHAUD-NETTO, J. and FOWLER, H.G., 2004. Body size and flight distance in stingless bees (Hymenoptera:meliponini): inference of flightrange and possible ecological implications. Brazilian Journal of Biology = Revista Brasileira de Biologia, vol. 64, no. 3B, pp. 563-568. http://dx.doi.org/10.1590/S1519-69842004000400003 PMid:15619994.
    » http://dx.doi.org/10.1590/S1519-69842004000400003
  • ASSEMI, H., SAJJADIE, A. and NAGHIZADEH, F., 2014. Investigation of different values of nano imidacloprid for control Tabaco aphid Myzus persicae nicotianae in Laboratory. Agrotechnology Journal., vol. 3, pp. 128. http://dx.doi.org/10.4172/2168-9881.1000128
    » http://dx.doi.org/10.4172/2168-9881.1000128
  • BLUM, M.S., FALES, H.M., TUCKER, K.W. and CPLLINS, A.M., 1978. Chemistry of the sting apparatus of theworker honeybee. Journal of Apicultural Research, vol. 17, no. 4, pp. 218-221. http://dx.doi.org/10.1080/00218839.1978.11099929
    » http://dx.doi.org/10.1080/00218839.1978.11099929
  • CALVERT, J.G., HEYWOOD, J.B., SAWYER, R.F. and SEINFELD, J.H., 1993. A chieving acceptable air quality: some reflections on controlling vehicle emissions. Science Journal., vol. 261, no. 5117, pp. 37-45. PMid:17750544.
  • CRESSWELL, J.E., DESNEUX, N. and VANENGELSDORP, D., 2012. Dietary traces of neonicotinoid pesticides as a cause of population declines in honey bees: An evaluation by Hill’s epidemiological criteria. Pest Management Science Journal, vol. 68, no. 6, pp. 819-827. http://dx.doi.org/10.1002/ps.3290 PMid:22488890.
    » http://dx.doi.org/10.1002/ps.3290
  • DACHER, M. and GAUTHIER, M., 2008. Involvement of NO-synthase and nicotinic receptors in learning in the honey bee. Physiological behavior Journal 95(1-2), 200-207.
  • DAINAT, B., EVANS, J.D., CHEN, Y.P., GAUTHIER, L. and NEUMANN, P., 2012. Predictive markers of honey bee colony collapse. PLoS One, vol. 7, no. 2, pp. e32151. http://dx.doi.org/10.1371/journal.pone.0032151 PMid:22384162.
    » http://dx.doi.org/10.1371/journal.pone.0032151
  • DAINAT, B., VANENGELSDORP, D. and NEUMANN, P., 2011. Colony collapse disorder in Europe. Environmental Microbiology Reports, vol. 4, no. 1, pp. 123-125. http://dx.doi.org/10.1111/j.1758-2229.2011.00312.x PMid:23757238.
    » http://dx.doi.org/10.1111/j.1758-2229.2011.00312.x
  • DECOURTYE, A., ARMENGAUD, C., RENOU, M., DEVILLERS, J., CLUZEAU, S., GAUTHIER, M. and PHAM-DELEGUE, M.H., 2004a. Imidacloprid impairs memory and brain metabolism in the honeybee (Apis mellifera L.). Pesticide Biochemistry and Physiology, vol. 78, no. 2, pp. 83-92. http://dx.doi.org/10.1016/j.pestbp.2003.10.001
    » http://dx.doi.org/10.1016/j.pestbp.2003.10.001
  • DECOURTYE, A., DEVILLERS, J., CLUZEAUl, S., CHARRETON, M., and PHAM-DELEGUE M.H., 2004b. Effects of Imidacloprid and deltamethrin on associative learning in honeybees under semifield and laboratory conditions. Ecotoxicology and Environmental Safety Journal, vol. 57, pp. 410-419.
  • DESNEUX, N., DECOURTYE, A. and DELPUECH, J.M., 2007. The sublethal effects of pesticides on beneficial arthropods. Annual Review of Entomology, vol. 52, no. 1, pp. 81-106. http://dx.doi.org/10.1146/annurev.ento.52.110405.091440 PMid:16842032.
    » http://dx.doi.org/10.1146/annurev.ento.52.110405.091440
  • DUKAS, R. and MORSE, D.H., 2003. Crab spiders affect flower visitation by bees. Oikos, vol. 101, no. 1, pp. 157-163. http://dx.doi.org/10.1034/j.1600-0706.2003.12143.x
    » http://dx.doi.org/10.1034/j.1600-0706.2003.12143.x
  • EFSA, 2012. EFSA panel on plant protection products and their residues (PPR); scientific opinion on the science behind the development of a risk assessment of plant protection products on bees (Apis mellifera, Bombus spp. and solitary bees). EFSA Journal, vol. 10, no. 5, pp. 2668. http://dx.doi.org/10.2903/j.efsa.2012.2668
    » http://dx.doi.org/10.2903/j.efsa.2012.2668
  • FINNEY, D., 1971. Probit analysis Cambridge, UK: Cambridge University Press, 256 p.
  • FISCHER, J., MÜLLER, T., SPATZ, A.K., GREGGERS, U., GRÜNEWALD, B. and MENZEL, R., 2014. Neonicotinoids interfere with specific components of navigation in honeybees. PLoS One, vol. 9, no. 3, pp. e91364. http://dx.doi.org/10.1371/journal.pone.0091364 PMid:24646521.
    » http://dx.doi.org/10.1371/journal.pone.0091364
  • FREE, J.B., 1987. Pheromones of social bees New York: Chapman & Hall, 236 p.
  • FREE, J.B., FERGUSON, A.W. and SIMPKINS, J.R., 1989. Honeybee responses to chemical components from the worker sting apparatus mandibular glands in field tests. Journal of Apicultural Research, vol. 28, no. 1, pp. 7-21. http://dx.doi.org/10.1080/00218839.1989.11100814
    » http://dx.doi.org/10.1080/00218839.1989.11100814
  • GAUTHIER, M. 2010. State of the art on insect nicotinic acetylcholine receptor function in learning and memory. In: S.H. THANY. Insect acetylcholine receptors New York: Springer, pp. 97-115. http://dx.doi.org/10.1007/978-1-4419-6445-8_9
    » http://dx.doi.org/10.1007/978-1-4419-6445-8_9
  • GOULSON, D., 2013. An overview of the environmental risks posed by neonicotinoid insecticides. Journal of Applied Ecology, vol. 50, no. 4, pp. 977-987. http://dx.doi.org/10.1111/1365-2664.12111
    » http://dx.doi.org/10.1111/1365-2664.12111
  • GRIFFITH, P.R. and FULLER, M.P., 1982. Advances in Infrared and Raman Spectroscopy Clark, R.J.H. and Hester, R. E., eds, Heydon & Sons: London, vol. 9, pp. 63-129.
  • GRIFFITH, P.R. and HASETH, J.A., 1986. Fourier transform infrared spectrometry New York: Wiley and Sons.
  • GUAN, H., CHI, D., YU, J. and LI, X., 2008. A novel photodegradable insecticide: Preparation, characterization and properties evaluation of nano-Imidacloprid. Pesticide Biochemistry and Physiology, vol. 92, no. 2, pp. 83-91. http://dx.doi.org/10.1016/j.pestbp.2008.06.008
    » http://dx.doi.org/10.1016/j.pestbp.2008.06.008
  • HAN, P., NIU, C. Y., LEI, C. L., CUI, J. J. and DESNEUX, N., 2010. Use of an innovative T-tube maze assay and the proboscis extension response assay to assess sublethal effects of GM products and pesticides on learning capacity of the honey bee Apis mellifera L. Ecotoxin Journal, vol. 19, pp. 1612-1619.
  • HARPAZ, I. and LENSKY, Y., 1959. Experiments on bee repellents. Bee World, vol. 40, no. 6, pp. 146-153. http://dx.doi.org/10.1080/0005772X.1959.11096718
    » http://dx.doi.org/10.1080/0005772X.1959.11096718
  • HENRY, M., BEGUIN, M., REQUIER, F., ROLLIN, O., ODOUX, J.F., AUPINEL, P., APATEL, J., TCHAMITCHIAN, S. and DECOURTYE, A., 2012. A common pesticide decreases foraging success and survival in honey bees. Science Journal, vol. 336, no. 6079, pp. 348-350. http://dx.doi.org/10.1126/science.1215039
    » http://dx.doi.org/10.1126/science.1215039
  • JOHNSON, J.D. and PETTIS, J.S., 2014. A survey of imidacloprid levels in water sources potentially frequented by honeybees (Apis mellifera) in the eastern USA. Water Air Soil Pollitten., vol. 225, no. 11, pp. 2127. http://dx.doi.org/10.1007/s11270-014-2127-2 PMid:25477698.
    » http://dx.doi.org/10.1007/s11270-014-2127-2
  • JOHNSON, R., PANKOW, J., BENDER, D., PRICE, C. and ZOGORISKI, J., 2000. MtbeT, to what extent will past releases contaminate community Water supply welly. Environmental Science Tecnology Journal, vol. 34, no. 9, pp. 210a-217a. http://dx.doi.org/10.1021/es003268z
    » http://dx.doi.org/10.1021/es003268z
  • JOHNSON, R.M., ELLIS, M.D., MULLIN, C.A. and FRAZIER, M., 2010. Pesticides and honey bee toxicity–USA. Apidologie, vol. 41, no. 3, pp. 312-331. http://dx.doi.org/10.1051/apido/2010018
    » http://dx.doi.org/10.1051/apido/2010018
  • JONES, J.C., HELLIWELL, P., BEEKMAN, M., MALESZKA, R. and OLDROYD, B.P., 2005. The effects of rearing temperature on developmental stability and learning and memory in the honey bee, Apis mellifera. Journal of Comparative Physiology, vol. 191, no. 12, pp. 1121-1129. http://dx.doi.org/10.1007/s00359-005-0035-z PMid:16049697.
    » http://dx.doi.org/10.1007/s00359-005-0035-z
  • KEVAN, P.G., 1999. Pollinators as bioindicators of the state of the environment; species, activity anddiversity. Agriculture Ecosystem Environmental Journal., vol. 74, no. 1-3, pp. 373-393. http://dx.doi.org/10.1016/S0167-8809(99)00044-4
    » http://dx.doi.org/10.1016/S0167-8809(99)00044-4
  • KEVAN, P.G., CLARK, E.A. and THOMAS, V.G., 1990. Pollination: A crucial ecological and mutualistic link inagro-forestry and sustainable agriculture. Proc Entomology Society Ont, vol. 121, pp. 43-48.
  • LAMBIN, M., ARMENGAUD, C., RAYMOND, S. and GAUTHIER, M., 2001. Imidacloprid‐induced facilitation of the proboscis extension reflex habituation in the honeybee. Archives of Insect Biochemistry and Physiology, vol. 48, no. 3, pp. 129-13. http://dx.doi.org/10.1002/arch.1065 PMid:11673842.
    » http://dx.doi.org/10.1002/arch.1065
  • MALERBO-SOUZA, D.T. and NOGUEIRA-COUTO, R.H., 2004. Efficiency of n-Octyl-Acetate, 2-Heptanone andcitronellal in repelling bees from basil (Ocimum sellowii – Labiatae). Brazilian Archives of Biology and Technology, vol. 47, no. 1, pp. 121-125. http://dx.doi.org/10.1590/S1516-89132004000100016
    » http://dx.doi.org/10.1590/S1516-89132004000100016
  • MAYER, D.F., 1997. Effects of methyl salicylate on honey bee (Apis mellifera L.) foraging. New Zealand Journal of Crop and Horticultural Science, vol. 25, no. 3, pp. 291-294. http://dx.doi.org/10.1080/01140671.1997.9514018
    » http://dx.doi.org/10.1080/01140671.1997.9514018
  • MENZEL, R., GREGGERS, U. and HAMMER, M., 1993. Functional organization of appetitive learning andmemory in a generalist pollinator, the honey bee. In: D.R. PAPAJ and A.C. LEWIS, eds. Insectlearning: Ecological and evolutionary perspectives New York: Chapman & Hall, pp. 79-125.
  • MICROSOFT EXCEL, 1997. Microsoft Excel Redmond, WA: Microsoft.
  • MORSE, R.A. and NOWOGRODZKI, R., 1990. Honey bee pests, predators, and diseases Ithaca, New York: Comstock Publishing Associates.
  • NATIONAL RESEARCH COUNCIL, 2007. Status of pollinators in North America Washington, DC: The National Academies Press, 322 pp
  • NAUEN, R., HUNGENBERG, H., TOLLO, B., TIETJEN, K. and ELBERT, A., 1998. Antifeedant effect, biologica l efficacy and high affinity binding of imidacloprid to acetylcholine receptors in Myzus persicae and Myzus nicotianae. Pesticides Science Journal, vol. 53, no. 2, pp. 133-140. http://dx.doi.org/10.1002/(SICI)1096-9063(199806)53:2<133::AID-PS756>3.0.CO;2-D
    » http://dx.doi.org/10.1002/(SICI)1096-9063(199806)53:2<133::AID-PS756>3.0.CO;2-D
  • PARVEEN, S.A., MARTIN, R.J. and SAHAYARAJ, K., 2014. Terminals Chebula Retz gallic acid – Based silver nanoparticles and their antiphytopathogenic activity. Journal of Biopesticides, vol. 7, pp. 1-6.
  • RAMIREZ-ROMERO, R., CHAUFAUX, J. and PHAM-DELEGUE, M.H., 2005. Effects of Cry1Ab protoxin, deltamethrin and imidacloprid on the foraging activity and the learning performances of the honeybee Apis mellifera, a comparative approach. Apidologie, vol. 36, no. 4, pp. 601-611. http://dx.doi.org/10.1051/apido:2005039
    » http://dx.doi.org/10.1051/apido:2005039
  • READER, T., HIGGINSON, A.D., BARNARD, C.J. and GILBERT, F.S., 2006. The effects of predation risk from crab spiders on bee foraging behavior. Behavior Ecology Journal., vol. 17, no. 6, pp. 933-939. http://dx.doi.org/10.1093/beheco/arl027
    » http://dx.doi.org/10.1093/beheco/arl027
  • REUTER, J.E., ALLEN, B.C., RICHARDS, R.C., PANKOW, J.F., GOLDMAN, C.R., SCHOLL, R.L. and SEYFRIED, J.S., 1998. Concentrations, souPerces, and fate of the gasoline oxygenate methyl/e/7-butyl ether (MTBE) in a multiple-use lake. Environmental Science & Technology, vol. 32, no. 23, pp. 3666-3672. http://dx.doi.org/10.1021/es9805223
    » http://dx.doi.org/10.1021/es9805223
  • RONDEAU, G., SÁNCHEZ-BAYO, F., TENNEKES, H.A., DECOURTYE, A., RAMÍREZ-ROMERO, R. and DESNEUX, N., 2014. Delayed and time-cumulative toxicity of imidacloprid in bees, ants and termites. Scientific Reports, vol. 4, pp. 1-8. PMid:24993452.
  • SABBOUR, M.M., 2015. Efficacy of some nano-Imidacloprid against red flour beetle Tribolium confusum (Coleoptera Tenebrionidae) under laboratory and stored conditions. Advances in Biochemistry & Biotechnology vol. 1, pp. 1-13.
  • SÁNCHEZ-BAYO, S., 2011. Insecticides Mode of Action in Relation to Their Toxicity to Non-Target Organisms. J Environment Analytic Toxicol, vol. S4. http://dx.doi.org/10.4172/2161-0525.S4-002
    » https://doi.org/10.4172/2161-0525.S4-002
  • SCHNEIDER, C.W., TAUTZ, J., GRÜNEWALD, B. and FUCHS, S., 2012. RFID tracking of sublethal effects of two neonicotinoid insecticides on the foraging behavior of Apis mellifera. PLoS One, vol. 7, no. 1, pp. e30023. http://dx.doi.org/10.1371/journal.pone.0030023 PMid:22253863.
    » http://dx.doi.org/10.1371/journal.pone.0030023
  • SILVA, F. S. and REBÊLO, J. M. M., 2002. Population dynamics of euglossinae bees(Hymenoptera, Apidae) in an earlysecond-growth forest of cajual island,in the state of Maranhão, Brazil. Brazilian Journal of Biology, vol. 62, 1, pp. 15-23.
  • SOKAL, R.R. and ROHLF, F.J., 1995. Biometry The Principles and Practice of Statistics in Biological Research. 3rd ed. New York: W.H. Freeman and Co.
  • SOLOMON, K.R., WILLIAMS, W.M., MACKAY, D., PURDY, J., GIDDINGS, J.M. and GIESY, J.P., 2014. Properties and uses of chlorpyrifos in the United States. Reviews of Environmental Contamination and Toxicology, vol. 231, pp. 13-34. http://dx.doi.org/10.1007/978-3-319-03865-0_2 PMid:24723132.
    » http://dx.doi.org/10.1007/978-3-319-03865-0_2
  • SURYANARAYANAN, S., 2013. Balancing control and complexity in field studies of neonicotinoids and honey bee health. Insects, vol. 4, no. 1, pp. 153-167. http://dx.doi.org/10.3390/insects4010153 PMid:26466800.
    » http://dx.doi.org/10.3390/insects4010153
  • VAN LEXMOND, M.B., BONMATIN, J.M., GOULSON, D. and NOOME, D.A., 2015. Worldwide integrated assessment on systemic pesticides: Global collapse of the entomofauna: Exploring the role of systemic insecticides. Environmental Science and Pollution Research, vol. 22, pp. 1-4.
  • VANENGELSDORP, D., HAYES, J., UNDERWOOD, R., CARON, D. and PETTIES, J., 2011. A survey of managed honey bee colony losses in the USA, Fall 2009 to Winter 2010. Journal of Apicultural Research, vol. 50, no. 1, pp. 1-10. http://dx.doi.org/10.3896/IBRA.1.50.1.01
    » http://dx.doi.org/10.3896/IBRA.1.50.1.01
  • WILLIAMSON, S.M. and WRIGHT, G.A., 2013. Exposure to multiple cholinergic pesticides impairs olfactory learning and memory in honeybees. The Journal of Experimental Biology, vol. 216, no. Pt 10, pp. 1799-1807. http://dx.doi.org/10.1242/jeb.083931 PMid:23393272.
    » http://dx.doi.org/10.1242/jeb.083931
  • WILSON, W.T. and MENAPACE, D.M., 1979. Disappearing disease of honey bees: A survey of the United States. American Bee Journal, vol. 119, pp. 184-186.
  • WU, Y.Y., ZHOU, T., WUBIE, A.J., WANG, Q., DAI, P.L. and JIA, H.R., 2014. Apoptosis in the nerve cells of adult honeybee (Apis mellifera ligustica) brain induced by imidacloprid. Acta Entomology Sinica, vol. 57, pp. 194-203.
  • YAMAMOTO, I., TOMIZAWA, M., SAITO, T., MIYAMOTO, T., WALCOTT, E.C. and SUMIKAWA, K., 1998. Structural factors contributing to insecticidal and selective actions of neonicotinoids. Archives of Insect Biochemistry and Physiological Journal, vol. 37, no. 1, pp. 24-32. http://dx.doi.org/10.1002/(SICI)1520-6327(1998)37:1<24::AID-ARCH4>3.0.CO;2-V PMid:9397513.
    » http://dx.doi.org/10.1002/(SICI)1520-6327(1998)37:1<24::AID-ARCH4>3.0.CO;2-V
  • ZHOU, T., SONG, H.L., WANG, Q., DAI, P.L., WU, Y.Y. and SUN, J.H., 2014. Effects of imidacloprid on the distribution of nicotine acetylcholine receptors in the brain of adult honeybee (Apis mellifera ligustica). Acta Entomology Sinica, vol. 56, pp. 1258-1266.

Publication Dates

  • Publication in this collection
    23 Oct 2020
  • Date of issue
    Oct-Dec 2021

History

  • Received
    18 Oct 2019
  • Accepted
    06 May 2020
  • Published
    30 Nov 2021
Instituto Internacional de Ecologia R. Bento Carlos, 750, 13560-660 São Carlos SP - Brasil, Tel. e Fax: (55 16) 3362-5400 - São Carlos - SP - Brazil
E-mail: bjb@bjb.com.br