DERLEME MAKALESİ / REVIEW ARTICLE IMPACT OF PESTICIDES ON HONEYBEE (Apis mellifera L.) DRONES Tarım İlaçlarının Bal Arılarında (Apis mellifera L.) Erkek Arı Üzerindeki Etkileri Faten BEN ABDELKADER Laboratory of Bioagressor and Integrated Protection in Agriculture, Department of Plant Health and Environment, National Institute of Agronomy of Tunisia (INAT), University of Carthage, Tunis, TUNISIA Bursa Uludağ Üniversitesi, Arıcılık Geliştirme Uygulama ve Araştirma Merkezi, 16059, Görükle Kampüsü Nilüfer/Bursa, TÜRKİYE, E-mail: benabdelkader.faten@gmail.com, ORCID No.: 0000-0003-4063-5521 Geliş Tarihi / Received:30.09.2019 Kabul Tarihi / Accepted: 23.10.2019 DOI: https://doi.org/10.31467/uluaricilik.626929 ABSTRACT Published research about drones is far less extensive than either worker or queen bees because they do not contribute to pollination, brood or honey production. However, much of the reproductive quality of the queen, though, is a function of the mating success and quality of the drones. Besides, studies of drones could help in breeding programs by improving the efficiency and quality of mating. Drones whose reproductive competitiveness is affected by several environmental and in-hive factors during development or adulthood may contribute dead or suboptimal sperm to a queen. It can have severe negative consequences not only for the queen herself but for overall productivity and survival of her colony. Drones are very sensitive to acaricides and insecticides. Most of them have negative impacts not only on drone semen quality such as spermatozoid viability and concentration but also on drone production and their traits. We here review the studies that describe pesticide exposure that might influence drone fitness. Key words: Drones, Acaricides, Insecticides, Fitness, Spermatozoa ÖZ Erkek arılarla yapılan yayınlar tozlaşma, yavru ve bal üretimine katkı sağlamadığı için işçi ve ana arılardan çok daha azdır. Fakat ana arının üremedeki başarısı kaliteli erkek arılarla başarılı bir şekilde çiftleşmesinin bir sonucudur. Bunun yanında erkek arı ile çalışmalar çiftleşmenin kalitesi ve etkinliğini artırabilir. Erkek arıların üremedeki rekabeti çevresel ve kovan içi gelişme faktörlerinden etkilenir ve bu durum ana arının ölümü veya ideal sperm seviyesinin altında kalmasına neden olabilir. Bu sorun ana arıyı olumsuz etkileme ve dolayısı ile koloninin toplam üretim ve yaşamını çok ciddi derecede olumsuz etkileyecek sonuçlar doğurabilir. Erkek arılar böcek ve akar öldürücü kimyasallara karşı çok hassastırlar. Bu kimysalların çoğu hem erkek arı sperm kalitesi, spermlerin yaşam gücü ve konsantrasyonu ve hem de erkek üretimi ve karakterleri üzerinde olumsuz etkileri bulunmaktadır. Bu ilaçlar erkek arıların yavru döneminde önce işçi arılar ve daha sonra hem işçi arılar ve hemde kendileri kovan içinde beslenirken olumsuz etkilenmektedir. Bu derleme çalışması tarım ilaçlarına maruz kalan erkek arıların nasıl etkilenebileceği konusundaki çalışmaları kapsamaktadır. Anahtar Kelimeler: Erkek arılar, Akarisit, Böcek öldürücüler, Uyum, Sperm Uludağ Arıcılık Dergisi – Uludag Bee Journal 2019, 19 (2): 188-194 188 DERLEME MAKALESİ / REVIEW ARTICLE GENIŞLETILMIŞ ÖZET Amaç: Bal arıları ekosistemdeki önemli rolleri ve son yıllarda bal arılarında koloni kayıpları nedeni ile bir çok çalışmanın konusu olmuştur. Bir bal arısı kolonisinde üremeden sorumlu bir ana arı olduğu için ilk akla gelen kolonideki ana arının durumudur. Bu yüzden koloninin geleceği ve üretkenliği için en önemli birey kolonide ana arıdır. Arıcılar ana arıları bu yüzden 1-2 yıl gibi bir sürede değitirmeye çalışırlar. Kolonilerin zayıflamasında ana arının yetersiz veya düşük kaliteli erkek arılar ile çiftleşmesi ana etkenlerden biridir. Tartışma: Erkek arılar ticari olarak önemli olmadığı ve kısıtlı bir zaman diliminde üretildiği için işçi arılar kadar yoğun olarak çalışılmamıştır. Erkek arılar zamanın çoğunu kovan içinde geçirsede özellikle larval dönemde ve daha sonra böcek öldrüücü veye tarım ilaçlarına maruz kalabilmektedir. Dolayısı ile erkek arılar hem arıcıların kovan içinde özellikle varroa parazitine karşı kullandığı ve çevredeki tarım ilaçlarından etkilenebilmektedir. Bu durumda varrao için kullanılan sentetik ilaçlar, organic asitler ve esansisyel yağlar gibi ilaçlar kullanılmaktadır. Bunun yanında bazı ülkelerde nosema ve yavru çürüklüğü hastalıkları için antibiyotikler de kullanılabilmektedir. Erkek arılarla yapılan yayınlar tozlaşma, yavru ve bal üretimine katkı sağlamadığı için işçi ve ana arılardan çok daha azdır. Fakat ana arının üremedeki başarısı kaliteli erkek arılarla başarılı bir şekilde çiftleşmesinin bir sonucudur. Bunun yanında erkek arı ile çalışmalar çiftleşmenin kalitesi ve etkinliğini artırabilir. Erkek arıların üremedeki rekabeti çevresel ve kovan içi gelişme faktörlerinden etkilenir ve bu durum ana arının ölümü veya ideal sperm seviyesinin altında kalmasına neden olabilir. Bu sorun ana arıyı olumsuz etkileme ve dolayısı ile koloninin toplam üretim ve yaşamını çok ciddi derecede olumsuz etkileyecek sonuçlar doğurabilir. Erkek arılar böcek ve akar öldürücü kimyasallara karşı çok hassastırlar. Bu kimysalların çoğu hem erkek arı sperm kalitesi, spermlerin yaşam gücü ve konsantrasyonu ve hem de erkek üretimi ve karakterleri üzerinde olumsuz etkileri bulunmaktadır. Bu ilaçlar erkek arıları yavru döneminde önce işçi arılar ve daha sonra hem işçi arılar ve hemde kendileri kovan içinde beslenirken olumsuz etkilemektedir. Sonuç: Tarım ilaçları ve kovan içinde başta varroa için kullanılan akar ve böcek öldürücü ilaçlar, organik asitler, esansiyel yağlar erkek arılarda önemli etkilere neden olmaktadır. Son yıllarda yapılan çalışmalarda yeni nesil tarım ilaçları olan neonikotinoidlerinde erkek arılar üzerinde olumsuz etkilere sahip olduğu rapor edilmiştir. Bu ilaçların erkek arıların yaşam süresi, doğum ağırlığı, kanat boyu, genişliği, uçuş faaliyetleri, sperm miktarı ve kalitesi üzerinde olumsuz etkileri olduğu yapılan çalışmalar sonucunda görülmektedir. Ayrıca bazı tarım ilaçları spermlerin yaşam gücünü ciddi derecede düşürmektedir. Bal arıları her ne kadar dişi merkezli canlılar olsada erkek arıların üremedeki başarısı doğal seleksiyon açısından ana etken olarak görülmekte olduğundan çevre faktörlerinin erkek arıların sağlığını nasıl etkilediğini belirlemek önemli bir konu olarak düşünülmektedir. INTRODUCTION (Winston 1987) and a principal focus of beekeepers. Honey bees, Apis mellifera L. (Hymenoptera: Although queens have a 3–4 year adult lifespan Apidae), play a highly complex and significant role in (Winston 1987), commercial beekeepers typically the ecosystem. They have been the focus of many replace their queens every 1–2 years because of the studies in recent years due to the progressive critical importance of a vigorous queen to colony decline in their number (Neumann and Carreck survival and productivity (Amiri et al. 2017). Multiple 2010, Potts et al. 2010, Goulson et al. 2015). drones are required to inseminate the queen with Pathogens and pesticides are thought to weaken the high-quality sperm on one or more mating flights immunity of bees and affect their physiology and (Woyke 1955). After the mating, the queen will store development (Frazier et al. 2008). To determine the the sperm in her spermathecae and fertilize eggs. main causes of the high rate of weakening in honey Queens generally mate with approximately 12–20 bee colonies, special attention has been focused on drones (Tarpy et al. 2015) and up to 45 drones queens (Traynor et al. 2016). The queen is the only (Neumann and Moritz 2000). Queens inseminated reproductive female of the colony and is the main with sub-fertile drones are themselves key when assessing colony reproductive output reproductively impaired. Thus, while “poor queens” may be a leading reason for colony failure, a major 189 U. Arı D. – U. Bee J. 2019, 19 (2): 188-194 DERLEME MAKALESİ / REVIEW ARTICLE cause of queens being perceived as poor is very Trophallaxis, the exchange of food by mouth, occurs likely to be “poor drones” (Kairo et al. 2016). among colony’s castes. Drones are fed by workers Drone Reproductive Development of all ages frequently (Haydak 1957). During their development, drone larvae receive more food than Compared to workers honeybee, drones have not worker larvae (avg. 9.6 mg versus 1.7 mg per cell) been thoroughly investigated because they are not (Haydak 1957). For the first few days of their lives, of direct commercial interest and they are reared drones are fed entirely by workers. Later, they are only during limited periods. Drones live less than both fed by workers and feed themselves from honey workers who live about 40–140 days depending on cells. The larval diet which is composed mainly of the season. Drones can live until 60 days (Page and pollen and nectar given to honey bee larvae expose Peng 2001) but some studies show that they can them transdermal, orally and internally; therefore, reach 90 days (Fukuda and Ohtani 1977). They the potential chronic toxicity and synergistic either die during mating or killed by workers when interactions at the brood stage seems likely to occur, the swarming season comes to an end. knowing that life stages might be much more The development of drones from egg stage until sensitive to certain contaminants compared to the emergence took approximately 24 days in larger adult stage (Zhu et al. 2014) cells compared to those of workers (Smith et al. Impact of Pesticides on Drone Fitness 2014). The reproductive biology of male honey bees is distinct from that of mammals. In drones, sperm The process of finding a queen and mate with her production starts in the testes during pupation and involves the capacity of drones to be able to reach a the sperm migrate to the seminal vesicles during the drone congregation area (DCA) (Koeniger et al. adult drone maturation, which is reached at least 16- 2014), locate the queen, compete with thousands of 18 days after adult drone emergence (Metz and other drones, and deliver sperm that the queen will Tarpy 2019). The migration of sperm to the seminal store in her spermatheca (Winston 1987). Hence, vesicles starts two days before eclosion. the ability to copulate which involves the body size Spermatozoa and the entire body of drones have to (Berg et al. 1997) and the ability to inseminate are undergo yet a physiological maturation process critical in order for a drone to offer a genetic diversity where the secretion of the glands of the genital tract passed to the next generation. is essential (Snodgrass 1984). Nutrition can affect Drones can be exposed to a large set of chemicals the timing of drone sexual maturation (Rhodes from their environment and from beekeeping 2008). Many factors (environmental and biotic) can practices. Agricultural activities can be a source of a also affect drone reproductive quality such as age, variety of pesticides. But also the products season, and genetics (Rhodes et al. 2010). administered to bee colonies by the beekeeper are Mature drones are most of the time in the hive. They at least equally noteworthy. Indeed, beehives are are either resting, feeding or cleaning themselves treated against the ectoparasitic mite Varroa (Fukuda and Ohtani 1977), and flights occur when destructor with many chemicals, like pyrethroid and conditions are favorable, around afternoon (Ruttner organophosphate acaricides. Also naturally 1966). Drones make on average 2–4 flights per day occurring organic acids and essential oils are used and fly up to 7 km from their hive. Orientation flights (Rosenkranz et al. 2010). Further, different last approximately 1–6 min, whereas mating flights antibiotics are commonly used in some countries for take about 20–30 min (Currie 1987). preventive or therapeutic treatment against American and European foulbrood, and against How Drones can Be Exposed to Pesticides? nosemosis (Reybroeck et al. 2012). Recent studies focused on the exposure ways of Synthetic compounds such as coumaphos (Check- pesticides on honeybee pollination services Mite™ or Perizin®), fluvalinate (Apistan®, Gabon®), (Goulson et al. 2015) with residues in nectar and flumethrin (Bayvarol®) and amitraz (Apivar®, pollen (Mullin et al. 2010), surface water (Wauchope Varidol®) are the main used chemicals to control V. 1978) and floral secretions and plant exudates destructor. In order to find more “natural” treatment (Girolami et al. 2009). Honeybees store these against varroasis, oxalic acid, formic acid products in the hive leading to exposure of brood and (Formidol®) and thymol (Thymovar®) have been hive products (Ravoet et al. 2015). introduced and are becoming increasingly used in organic beekeeping. Uludağ Arıcılık Dergisi – Uludag Bee Journal 2019, 19 (2): 188-194 190 DERLEME MAKALESİ / REVIEW ARTICLE Few studies focused on the impact of pesticides drone sperm number was lower in all treated drones traits such as body weight and wings length colonies. and width of drones. Rinderer et al. (1999) were the first to study the effect of fluvalinate (the active Fisher and Rangel (2018) investigated the effects of ingredient in the product Apistan®) on drone the drone-rearing beeswax exposed to miticides production. They found that mortality was higher in (amitraz with a mix of coumaphos and fluvalinate) fluvalinate-treated colonies (66.9%) in drones aged and agrochemicals (chlorpyrifos and chlorothalonil) between 12 and 18 days old, compared to control. found in hives. Mean sperm viability was significantly They also reported that surviving drones from lower in drones reread in all treated beeswax. They colonies treated with fluvalinate caused drones to also found that sexual maturity of treated drones have about 5% reduction in body weight and the groups reached when they were aged between 16 to interaction of Varroa and fluvalinate led to 10% 18 days of age. reduction of drone body weight. Drones from Shoukry et al. (2013) also evaluated the effects of coumaphos treated colonies presented lower two acaricides (fluvalinate, amitraz), two organic viability in the first week (86%) compared to drones acids (oxalic acid, formic acid) and thymol on drone from untreated colonies (90%). The viability reached fertility. The lowest sperm number was found in the lowest rate by week 6, with a percentage of 49% drones exposed to fluvalinate and amitraz, while a compared to control (85%) (Burley et al. 2008). concordance was observed between sperm number Shoukry et al. (2013), treated emerged drones with and wing length among all treatment groups. different miticides. They found a reduction in wing length ranged from 5.36% in drones treated with Besides acaricides, several agrochemicals have fluvalinate compared to control drones. Also, been found to negatively affect the drone semen fluvalinate and amitraz treatments significantly quality. Oral exposure to the neonicotinoid decreased the wing width of drones by insecticides such as thiamethoxam and clothianidin approximately 4.57% and 2.27% respectively are known to reduce sperm viability in adult drones compared to the control. (Straub et al. 2016). Chronic exposure of sublethal doses of clothianidin at sexual maturity stage The impact of organic treatments on drone survival decreased semen volume and its concentration and and traits were also investigated by De Guzman et increased sperm mortality rate (Ben Abdelkader et al. (1999). Treated colonies with formic acid al. 2018). It also induced oxidative stress in produced less than half of drones than untreated spermatozoa by increasing antioxidants enzymes colonies. A reduction of the survival of ten days old (superoxide dismutase, glutathione peroxidase, and drones and a reduction of wing length were also catalase) and malondialdehyde level, which is a lipid reported. Drones treated with formic acid and oxalic peroxidation marker, in spermatozoa of drones acid induced a reduction in wing length and a exposed and also decreasing the protein content in reduction of 2.31% and 4.52%, respectively semen (Ben Abdelkader et al. 2019). (Shoukry et al. 2013). Furthermore, a high percentage of oxalic acid (more than 0.5%) reduced Exposure to imidacloprid was also found to affect the the survival of the drones (Aboushaara et al. 2017). mitochondrial activity of spermatozoa and therefore Sublethal doses of thymol treatment caused a the sperm viability (Ciereszko et al. 2017). reduction in drone flight activity (Johnson et al. Phynelperazole insecticides also affected the drone 2013). reproductive system. Sublethal doses of fipronil at sexual maturity led to a decrease of spermatozoa Given the reproductive quality of drones, most of the concentration, sperm viability and an increase in studies were focused on the impact of acaricides and sperm metabolic rate (Kairo et al. 2016). The co- insecticides on drone fertility and semen quality. In exposure of fipronil with the microsporidian parasite fact, coumaphos caused reduced sperm viability Nosema ceranae induced metabolic disturbances immediately after semen collection and in samples and affect oxidative stress defense in sperm. stored up to 6 weeks (De Guzman et al. 1999). Burley (2007) explored the effects of Apistan® Moreover, the acute in vitro exposure of six (fluvalinate), Checkmite+® (coumaphos), or Apilife molecules (fipronil, ethiprole, imidacloprid, Var® on sperm number and viability in the seminal thiamethoxam, cypermethrin, and coumaphos) of vesicles of mature drones. The lower sperm viability spermatozoid at different concentrations ranged was found in coumaphos treated colonies. Besides, from 0.1 to 100 µM led to an increase of 191 U. Arı D. – U. Bee J. 2019, 19 (2): 188-194 DERLEME MAKALESİ / REVIEW ARTICLE spermatozoid ATP levels. Fipronil, ethiprole, Apicultural Research DOI: imidacloprid and thiamethoxam significantly 10.1080/00218839.2019.1655182: 1-6. decreased the viability of spermatozoids (Ben Ben Abdelkader F., Kairo G., Tchamitchian S., Abdelkader et al. 2015) in 24 hours. Bonnet M., Cousin M., Barbouche N., Belzunces L., Brunet J. 2018. Effects of clothianidin exposure on semen parameters CONCLUSION of honey bee drones. Journal of New Managed honey bee colonies are exposed to Sciences 59: 3791-3798. multiple pesticides including insecticides such as Berg S., Koeniger N., Koeniger G., Fuchs S. 1997. neonicotinoids widely used in the foraging Body size and reproductive success of environment or miticides used to drones (Apis mellifera L). Apidologie 28(6): control Varroa mites, which are still widespread in 449-460. hive products and larvae food, can negatively impact Burley LM., Fell RD., Saacke RG. 2008. Survival of drone fitness. 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