Biochemical and physiological parameters of honey bee (Apis mellifera L.) during summer and winter in habitats with different anthropogenic influence

Medonosna pčela (Apis mellifera  Linnaeus, 1758) je široko rasprostranjena vrsta iz grupe socijalnih insekata, glavni oprašivač mnogih divljih biljaka i poljoprivrednih kultura. Pored njene primarne uloge u oprašivanju, koristi se i za dobijanje pčelinjih proizvoda, pre svega meda, voska  i propolisa,  čime se dodatno naglašava njen privredni značaj. Uzimajući u obzir značaj medonosne pčele, a sa druge strane aktuelan problem pada broja kolonija, postoji potreba za razumevanje uzroka  i  procenu  rizika koji leže u osnovi ovog kompleksnog problema. Stoga, istraživanja u ovoj doktorskoj disertaciji obuhvataju praćenje biohemijskih i fizioloških parametara kod medonosne pčele kao pokazatelja stanja pčelinjih zajednica tokom letnjeg i zimskog perioda  na staništima sa različitim antropogenim uticajem.  U prvoj fazi istraživanja su izvedeni akutni laboratorijski testovi izlaganja  medonosne pčele  subletalnim dozama pesticida, tiakloprida i klotianidina, a dobijeni rezultati su  ukazali  na narušenu neurotransmisiju, na izmene u aktivnosti antioksidativnih enzima i redoks statusa u ćeliji, kao i na imunosupresiju.  U drugoj fazi istraživanja su analizirane kolonije  medonosne pčele sa različitih lokaliteta  na području AP Vojvodine, sa primenom stacionarnog i migratornog tipa pčelarenja, tokom letnjeg i zimskog perioda, a dobijeni rezultati su ukazali  na značajne razlike  u biohemijskom odgovoru medonosne pčele u zavisnosti od starosti pčela, sezone godine i tipa pčelinjaka. Na osnovu rezultata prethodne dve faze istraživanja, izvršen  je  odabir uzoraka, kao i izbor parametara za  treću fazu  istraživanja,  gde  je praćeno stanje medonosne pčele  iz stacionalnih pčelinjih zajednica, tokom letnjeg  aktivnog perioda, kao i početkom  i  krajem  zimskog perioda  mirovanja pčela, sa tri lokaliteta u Republici Srbiji sa različitim stepenom urbanizacije i industrijalizacije: Beograd kao urbano, Susek kao ruralno i Zajača kao industrijsko područje. Dobijeni rezultati su ukazali na  značajne razlike u zavisnosti od sezone  godine, kao i  lokaliteta sa različitim antropogenim uticajem.  Zimske pčele, krajem perioda mirovanja  u odnosu na početak mirovanja,  karakteriše viši  nivo antioksidativne zaštite, udružen sa  visokim  stepenom  oksidativnih oštećenja, zatim  bolja imunokompetencija, kao i niske energetske rezerve u masnom telu.  Sezonskim povećanjem temperatura tokom godine povećava se sadržaj zasićenih, dok se smanjuje udeo nezasićenih masnih kiselina, gde se oleinska 18:1(9) kiselina izdvaja kao dominantna komponenta u ukupnom sadržaju masnih kiselina masnog tela  pčela  sa sve tri lokacije, tokom sva tri perioda.  Niži nivo ukupnih ugljovodonika je još jedna fiziološka karakteristika zimskih pčela. Takođe, zapažene su promene  i  u neurotransmisiji holinergičkog tipa tokom sezone, sa višim aktivnostima acetilholinesteraze kod pčela krajem zime i tokom leta.  Metabolomičkom analizom hemolimfe pčela je  identifikovano  125 metabolita, uključenih u 36 metaboličkih puteva, gde se kao biomarkeri od značaja za razlikovanje letnjih i zimskih pčela izdvajaju  metaboliti u metabolizmu glicerolipida i aminokiseline glicin, cistein, glutamin, fenilalanin, prolin i lizin.  Poređenjem dobijenih rezultata između različitih lokaliteta,  pčele iz Beograda se izdvajaju sa nižim nivoom oksidativnog stresa,  nižim aktivnostimaacetilholinesteraze, kao i fenol oksidaze i glukoza oksidaze, ključnih enzima u imunom odgovoru pčela. Pčele iz Suseka su se izdvojile sa najvećom masom masnog tela, što ukazuje na dobre nutritivne i energetske rezerve u pčelama tokom zime i leta, dok se Beograd izdvaja sa značajno višim udelom nezasićenih masnih kiselina  u masnom telu pčela.  Takođe su zabeležene razlike u relativnoj zastupljenosti  n-alkana u pčelama sa različitih lokaliteta, te je u Beogradu najzastupljeniji nonakozan  (C29), dok je heptakozan (C27) najzastupljeniji u uzorcima pčela iz Suseka i Zajače.  Analizom sadržaja metala u pčelama, pergi i medu, zapažamo da svaku sredinu karakteriše specifičan profil zagađenja,  kao i da je u medu znatno niža koncentracija svih analiziranih metala.  Dobijeni rezultati pružaju dobru osnovu za dalja ispitivanja biološkog odgovora medonosne pčele, u pogledu uticaja faktora okruženja, a time i dalje definisanje odgovarajućih mera zaštite pčelinjih zajednica.The  honey  bee  (Apis  mellifera  Linnaeus,  1758)  is  a‚widespread  social  insect  species  and  the  main pollinator of various wild plants and agricultural crops.In addition to its primary role in pollination, the honey bee  is  also  used  for  obtaining  honeybee  products, mostly honey, wax, and propolis, with a significant role in  the  economy.  Number  of  honey  bee  colonies  is declining,  thus  necessitating  the  assessment  of underlying  reasons  and  risks.  Therefore,  in  this doctoral  dissertation,  biochemical  and  physiological parameters  in  the  honey  bee  were  monitored  as indicators  of  the  colonies’  condition  during  the summer  and  winter  period  in  habitats  with  different anthropogenic  impacts.  In  the  first  phase  of  the research,  acute laboratory tests of honey bee exposure to  sublethal  doses  of  pesticides  (thiacloprid  and clothianidin)  were performed and the obtained results indicated  impaired  neurotransmission,  changes  in  the activity of antioxidant enzymes, and redox status in the cell,  as  well  as  immunosuppression.  In  the  second phase  of  the  research,  honey  bee  colonies  from different  localities  in  the  area  of  AP  Vojvodina  were analyzed,  using  the  stationary  and  migratory  types  of beekeeping, during the summer and winter period, and the obtained results indicated significant differences in the  biochemical response of the honey bee depending on the age of the bees, the season of the year, and the type of apiary. Based on the results of the previous two phases  of  the  research,  the  selection  of  samples  was performed,  as  well  as  the  selection  of  parameters  for analysis within the third phase of the research, where the  condition  of  honey  bees  from  stationary  bee communities was monitored during the summer active period,  as  well  as  the  beginning  and  end  of  winter dormancy of bees, from three localities in the Republic of  Serbia  with  different  degrees  of  urbanization  and industrialization: Belgrade as urban, Susek as rural and Zajaca  as  industrial  area.  The  obtained  results indicated  significant  differences  depending  on  the season  of  the  year,  as  well  as  the  location  with different  anthropogenic  influence.  Older  winter  bees are  characterized  by  a  higher  level  of  antioxidant protection,  associated  with  a  high  degree  of  oxidative damage,  better  immunocompetence,  as  well  as  lowenergy reserves in the fat body. The seasonal increase in  the  temperature  during  the  year  increases  the content  of  saturated  while  decreasing  the  ratio  of unsaturated  fatty  acids,  where  oleic  18:1(9)  acid stands  out  as  the  dominant  component  in  the  total fatty  acid  content  of  bee  body  fat  from  all  three locations,  during  all  three  periods.  The  lower  level  of total  hydrocarbons  is  another  physiological characteristic  of  winter  bees.  Also,  changes  in cholinergic  neurotransmission  were  observed  during the  season,  with  higher  acetylcholinesterase  activities in  bees  in  late  winter  and  summer.  Metabolomic analysis of bee hemolymph identified 125 metabolites,included in 36 metabolic pathways, where metabolites in  the  metabolism  of  glycerolipids  and  amino  acids glycine,  cysteine,  glutamine,  phenylalanine,  proline, and  lysine  stand  out  as  biomarkers  of  importance  for distinguishing summer and winter bees. By  comparing the obtained results between different localities, honey bees  from  Belgrade  stand  out  with  lower  levels  of oxidative  stress,  lower  activities  of acetylcholinesterase  as  well  as  phenoloxidase  and glucose oxidase, key enzymes in the immune response of  bees.  The  honey  bees  from  Susek  had  the  largest mass of the fat body, which indicates good nutritional and  energy  reserves  in  bees   during  winter  and summer,  while  the  honey  bees  from  Belgrade  were shown  to  have  a  significantly  higher  ratio  of unsaturated  fatty  acids  in  the  fat  body  of  bees  during the  year.  There  were  also  differences  in  the  relative prevalence  of  n-alkanes  in  honey  bees  from  different localities, nonacosane (C29) being the most common in Belgrade, while heptacosane (C27) is most prevalent in honey  bee  samples  from  Susek  and  Zajaca.  By analyzing the metal content in honey bees, perga, and honey,  it  was  noted  that  each  environment  is characterized by a specific pollution profile, as well as that the concentration of all analyzed metals in honey is significantly lower compared to samples of bees and perga.  The  obtained  results  provide  a  good  basis  for further studies of the biological response of  the honey bee,  in  terms  of  the  influence  of  the  environmental factors,  enabling  further  defining  of  appropriate measures for the protection of honey bee colonies

Biochemical and physiological parameters of honey bee (Apis mellifera L.) during summer and winter in habitats with different anthropogenic influence

Medonosna pčela (Apis mellifera  Linnaeus, 1758) je široko rasprostranjena vrsta iz grupe socijalnih insekata, glavni oprašivač mnogih divljih biljaka i poljoprivrednih kultura. Pored njene primarne uloge u oprašivanju, koristi se i za dobijanje pčelinjih proizvoda, pre svega meda, voska  i propolisa,  čime se dodatno naglašava njen privredni značaj. Uzimajući u obzir značaj medonosne pčele, a sa druge strane aktuelan problem pada broja kolonija, postoji potreba za razumevanje uzroka  i  procenu  rizika koji leže u osnovi ovog kompleksnog problema. Stoga, istraživanja u ovoj doktorskoj disertaciji obuhvataju praćenje biohemijskih i fizioloških parametara kod medonosne pčele kao pokazatelja stanja pčelinjih zajednica tokom letnjeg i zimskog perioda  na staništima sa različitim antropogenim uticajem.  U prvoj fazi istraživanja su izvedeni akutni laboratorijski testovi izlaganja  medonosne pčele  subletalnim dozama pesticida, tiakloprida i klotianidina, a dobijeni rezultati su  ukazali  na narušenu neurotransmisiju, na izmene u aktivnosti antioksidativnih enzima i redoks statusa u ćeliji, kao i na imunosupresiju.  U drugoj fazi istraživanja su analizirane kolonije  medonosne pčele sa različitih lokaliteta  na području AP Vojvodine, sa primenom stacionarnog i migratornog tipa pčelarenja, tokom letnjeg i zimskog perioda, a dobijeni rezultati su ukazali  na značajne razlike  u biohemijskom odgovoru medonosne pčele u zavisnosti od starosti pčela, sezone godine i tipa pčelinjaka. Na osnovu rezultata prethodne dve faze istraživanja, izvršen  je  odabir uzoraka, kao i izbor parametara za  treću fazu  istraživanja,  gde  je praćeno stanje medonosne pčele  iz stacionalnih pčelinjih zajednica, tokom letnjeg  aktivnog perioda, kao i početkom  i  krajem  zimskog perioda  mirovanja pčela, sa tri lokaliteta u Republici Srbiji sa različitim stepenom urbanizacije i industrijalizacije: Beograd kao urbano, Susek kao ruralno i Zajača kao industrijsko područje. Dobijeni rezultati su ukazali na  značajne razlike u zavisnosti od sezone  godine, kao i  lokaliteta sa različitim antropogenim uticajem.  Zimske pčele, krajem perioda mirovanja  u odnosu na početak mirovanja,  karakteriše viši  nivo antioksidativne zaštite, udružen sa  visokim  stepenom  oksidativnih oštećenja, zatim  bolja imunokompetencija, kao i niske energetske rezerve u masnom telu.  Sezonskim povećanjem temperatura tokom godine povećava se sadržaj zasićenih, dok se smanjuje udeo nezasićenih masnih kiselina, gde se oleinska 18:1(9) kiselina izdvaja kao dominantna komponenta u ukupnom sadržaju masnih kiselina masnog tela  pčela  sa sve tri lokacije, tokom sva tri perioda.  Niži nivo ukupnih ugljovodonika je još jedna fiziološka karakteristika zimskih pčela. Takođe, zapažene su promene  i  u neurotransmisiji holinergičkog tipa tokom sezone, sa višim aktivnostima acetilholinesteraze kod pčela krajem zime i tokom leta.  Metabolomičkom analizom hemolimfe pčela je  identifikovano  125 metabolita, uključenih u 36 metaboličkih puteva, gde se kao biomarkeri od značaja za razlikovanje letnjih i zimskih pčela izdvajaju  metaboliti u metabolizmu glicerolipida i aminokiseline glicin, cistein, glutamin, fenilalanin, prolin i lizin.  Poređenjem dobijenih rezultata između različitih lokaliteta,  pčele iz Beograda se izdvajaju sa nižim nivoom oksidativnog stresa,  nižim aktivnostimaacetilholinesteraze, kao i fenol oksidaze i glukoza oksidaze, ključnih enzima u imunom odgovoru pčela. Pčele iz Suseka su se izdvojile sa najvećom masom masnog tela, što ukazuje na dobre nutritivne i energetske rezerve u pčelama tokom zime i leta, dok se Beograd izdvaja sa značajno višim udelom nezasićenih masnih kiselina  u masnom telu pčela.  Takođe su zabeležene razlike u relativnoj zastupljenosti  n-alkana u pčelama sa različitih lokaliteta, te je u Beogradu najzastupljeniji nonakozan  (C29), dok je heptakozan (C27) najzastupljeniji u uzorcima pčela iz Suseka i Zajače.  Analizom sadržaja metala u pčelama, pergi i medu, zapažamo da svaku sredinu karakteriše specifičan profil zagađenja,  kao i da je u medu znatno niža koncentracija svih analiziranih metala.  Dobijeni rezultati pružaju dobru osnovu za dalja ispitivanja biološkog odgovora medonosne pčele, u pogledu uticaja faktora okruženja, a time i dalje definisanje odgovarajućih mera zaštite pčelinjih zajednica.The  honey  bee  (Apis  mellifera  Linnaeus,  1758)  is  a‚widespread  social  insect  species  and  the  main pollinator of various wild plants and agricultural crops.In addition to its primary role in pollination, the honey bee  is  also  used  for  obtaining  honeybee  products, mostly honey, wax, and propolis, with a significant role in  the  economy.  Number  of  honey  bee  colonies  is declining,  thus  necessitating  the  assessment  of underlying  reasons  and  risks.  Therefore,  in  this doctoral  dissertation,  biochemical  and  physiological parameters  in  the  honey  bee  were  monitored  as indicators  of  the  colonies’  condition  during  the summer  and  winter  period  in  habitats  with  different anthropogenic  impacts.  In  the  first  phase  of  the research,  acute laboratory tests of honey bee exposure to  sublethal  doses  of  pesticides  (thiacloprid  and clothianidin)  were performed and the obtained results indicated  impaired  neurotransmission,  changes  in  the activity of antioxidant enzymes, and redox status in the cell,  as  well  as  immunosuppression.  In  the  second phase  of  the  research,  honey  bee  colonies  from different  localities  in  the  area  of  AP  Vojvodina  were analyzed,  using  the  stationary  and  migratory  types  of beekeeping, during the summer and winter period, and the obtained results indicated significant differences in the  biochemical response of the honey bee depending on the age of the bees, the season of the year, and the type of apiary. Based on the results of the previous two phases  of  the  research,  the  selection  of  samples  was performed,  as  well  as  the  selection  of  parameters  for analysis within the third phase of the research, where the  condition  of  honey  bees  from  stationary  bee communities was monitored during the summer active period,  as  well  as  the  beginning  and  end  of  winter dormancy of bees, from three localities in the Republic of  Serbia  with  different  degrees  of  urbanization  and industrialization: Belgrade as urban, Susek as rural and Zajaca  as  industrial  area.  The  obtained  results indicated  significant  differences  depending  on  the season  of  the  year,  as  well  as  the  location  with different  anthropogenic  influence.  Older  winter  bees are  characterized  by  a  higher  level  of  antioxidant protection,  associated  with  a  high  degree  of  oxidative damage,  better  immunocompetence,  as  well  as  lowenergy reserves in the fat body. The seasonal increase in  the  temperature  during  the  year  increases  the content  of  saturated  while  decreasing  the  ratio  of unsaturated  fatty  acids,  where  oleic  18:1(9)  acid stands  out  as  the  dominant  component  in  the  total fatty  acid  content  of  bee  body  fat  from  all  three locations,  during  all  three  periods.  The  lower  level  of total  hydrocarbons  is  another  physiological characteristic  of  winter  bees.  Also,  changes  in cholinergic  neurotransmission  were  observed  during the  season,  with  higher  acetylcholinesterase  activities in  bees  in  late  winter  and  summer.  Metabolomic analysis of bee hemolymph identified 125 metabolites,included in 36 metabolic pathways, where metabolites in  the  metabolism  of  glycerolipids  and  amino  acids glycine,  cysteine,  glutamine,  phenylalanine,  proline, and  lysine  stand  out  as  biomarkers  of  importance  for distinguishing summer and winter bees. By  comparing the obtained results between different localities, honey bees  from  Belgrade  stand  out  with  lower  levels  of oxidative  stress,  lower  activities  of acetylcholinesterase  as  well  as  phenoloxidase  and glucose oxidase, key enzymes in the immune response of  bees.  The  honey  bees  from  Susek  had  the  largest mass of the fat body, which indicates good nutritional and  energy  reserves  in  bees   during  winter  and summer,  while  the  honey  bees  from  Belgrade  were shown  to  have  a  significantly  higher  ratio  of unsaturated  fatty  acids  in  the  fat  body  of  bees  during the  year.  There  were  also  differences  in  the  relative prevalence  of  n-alkanes  in  honey  bees  from  different localities, nonacosane (C29) being the most common in Belgrade, while heptacosane (C27) is most prevalent in honey  bee  samples  from  Susek  and  Zajaca.  By analyzing the metal content in honey bees, perga, and honey,  it  was  noted  that  each  environment  is characterized by a specific pollution profile, as well as that the concentration of all analyzed metals in honey is significantly lower compared to samples of bees and perga.  The  obtained  results  provide  a  good  basis  for further studies of the biological response of  the honey bee,  in  terms  of  the  influence  of  the  environmental factors,  enabling  further  defining  of  appropriate measures for the protection of honey bee colonies

Determination of residues of sulfonylurea herbicides in soil by using microwave-assisted extraction and high performance liquid chromatographic method

A modified method for the analysis of nicosulfuron, rimsulfuron and prosulfuron was developed and validated by using microwave-assisted extraction (MAE) and ultra-performance liquid chromatography with diode array detection in the ultraviolet region (HPLC-UV-DAD). The most important experimental parameters of extraction procedure and HPLC-UV-DAD technique were optimised in respect to those sulfonylurea herbicides. High recoveries of the microwave-assisted extraction were obtained by using a dichloromethane-acetonitrile mixture (2:1 volume ratio) acidified with acetic acid (0.8 vol.%) with the addition of urea. The mean recoveries at three spiking levels ranged from 97.47 to 98.76% for nicosulfuron, 97.88 to 99.17% for rimsulfuron and from 97.91 to 99.83% for prosulfuron. The limits of detection of nicosulfuron, rimsulfuron and prosulfuron were 0.95, 0.91 and 0.89 mu g kg(-1), respectively. The accuracy of the developed method was confirmed by HPLC coupled with tandem mass spectrometry parallel analyses. The developed method was used to investigate the dissipation dynamics of sulfonylurea herbicides in the real field trials in Vojvodina Province, Serbia. The obtained half-lives were 0.05, 0.23 and 0.15 days for recommended dose application of nicosulfuron, rimsulfuron and prosulfuron, respectively. Low residues and short half-life in soil suggested that the risk to sensitive rotational crops after application of those sulfonylurea herbicides is low when they are used in the appropriate dosages

The influence of low temperature and diapause phase on sugar and polyol content in the European corn borer Ostrinia nubilalis (Hbn.).

The European corn borer (ECB, Ostrinia nubilalis Hbn.) is a major pest in temperate regions of Europe and North America. Fifth instar ECB larvae enter diapause before winter and gradually develop cold hardiness. Here we investigated the combined influence of diapause phase and low temperature on sugar and polyol content in ECB larvae. Larvae in mid-diapause or diapause termination were acclimated at 5 °C, -3°C or -16 °C, and sugar and polyol content was measured using GC-MS. Control GC-MS measurements were conducted on untreated non-diapausing larvae. We detected differences in polyol (glycerol, sorbitol, myo-inositol) and sugar (trehalose, fructose, glucose) levels in diapausing versus non-diapausing larvae. Glycerol and trehalose were the most abundant of all analyzed cryoprotective compounds in diapausing larvae. Exposure of diapausing larvae to decreasing temperatures induced changes in polyol and sugar levels that depended on the phase of diapause. In mid-diapause larvae, decreasing temperatures induced a significant increase in glycerol and a decrease in sorbitol and myo-inositol. In larvae at diapause termination, polyol content was lower and less influenced by decreasing temperatures. In contrast, sugar levels were lower in larvae at mid-diapause versus diapause termination. Exposure of larvae to -16 °C induced a significant increase in the levels of all detected sugars. In particular, glucose levels were significantly higher in larvae at diapause termination following exposure to -16 °C. We propose that this shift toward sugar synthesis following low temperature exposure in larvae at diapause termination is a consequence of NADPH dependent polyol synthesis, and may be a mechanism for preservation of carbon reserves needed for post-diapause development

Laboratory bioassays on the response of honey bee (Apis mellifera L.) glutathione S-transferase and acetylcholinesterase to the oral exposure to copper, cadmium, and lead.

In the present study, the influence of cadmium, copper, and lead on two enzymes often used as biomarkers in toxicological analysis was investigated. Bees were fed with 1 M sucrose solution containing 10-fold serial dilutions of CuCl2 (1000 mg L-1, 100 mg L-1, and 10 mg L-1), CdCl2 (0.1 mg L-1, 0.01 mg L-1, and 0.001 mg L-1), or PbCl2 (10 mg L-1, 1 mg L-1, and 0.1 mg L-1) during 48 h. Our results showed that the total glutathione S-transferase activity was not changed under the influence of cadmium and lead, and it was decreased with the highest concentration of copper. The level of gene expression of the three analyzed classes of glutathione S-transferase was significantly increased with increasing concentrations of copper and cadmium. Lead did not cause significant changes in glutathione S-transferase activity and gene expression, while it showed biphasic effect on acetylcholinesterase activity: lower concentration of lead, 0.1 mg L-1 inhibited and higher dose, 10 mg L-1 induced acetylcholinesterase activity in honey bees. Furthermore, our results showed a significant decrease of the acetylcholinesterase activity in honey bees treated with 0.001 and 0.01 mg L-1 CdCl2. Our results indicate the influence of cadmium, copper, and lead on GST and AChE in the honey bees. These results form the basis for future research on the impact of metallic trace element pollution on honey bees

Exposure to high concentrations of cadmium which delay development of Ostrinia nubilalis Hbn. larvae affected the balance of bioelements

All processes involved in metal homeostasis must be coordinated to provide sufficient, but not toxic, concentrations of important bioelements, and to minimize detrimental effects of toxic metals. Our previous studies dealing with the exposure of O. nubilalis non-diapausing larvae to dietary Cd demonstrated that exposure to higher concentrations of Cd caused delay in the development of larvae, induced oxidative stress and also induced defense mechanisms against the toxic effects of Cd. The aim of the present study was to evaluate how acute and chronic exposure of O. nubilalis larvae to increased concentrations of dietary Cd affected the balance of important bioelements. The concentration of bioelements was analyzed in larvae (after short-term exposure) and pupae (after long-term exposure). The short-term exposure of final instar larvae (L5) to Cd did not affect significantly the concentration of any of the analyzed bioelements, while the long-term exposure of developing larvae to higher concentrations of Cd caused increase in the concentrations of Ca, Mg and Na in pupae. The bioaccumulation factor, calculated for bioelements after long-term exposure to Cd, was higher for the most bioelements in groups fed with diet containing higher concentrations of Cd, except K which displayed the opposite trend. Pearson correlation coefficient showed positive correlations between Cd and Ca, Mg, Na, Fe, Cu and Zn, while negative correlation was observed between Cd and K. The results indicate that impact on the balance of important bioelements might be one of the mechanisms of cadmium toxicity and certainly raise numerous questions for future research

The impact of sublethal concentrations of Cu, Pb and Cd on honey bee redox status, superoxide dismutase and catalase in laboratory conditions.

In this study, laboratory bioassays were performed to investigate the impact of sublethal concentrations of Cu (CuCl2: 1000, 100, 10 mg L(-1)), Pb (PbCl2: 10, 1, 0.1 mg L(-1)) and Cd (CdCl2: 0.1, 0.01, 0.001 mg L(-1)) on honey bee redox status and the activity of the main antioxidative enzymes and their gene expression. Our results show that exposure to these metals led to significant changes of gene expression, the levels of enzyme activity and redox status, but the effects are metal and dose dependent. In general, exposure of 48 h to given concentrations of Cu, Cd and Pb did not change the activity of antioxidative enzymes and the level of lipid peroxidation, with the exception of decreased activity of catalase at the lowest concentration of cadmium. Only lead produced increases in glutathione and thiol groups. Expression of genes for catalase and superoxide dismutase changed with exposure to cadmium and copper, whilst lead induced only expression of superoxide dismutase genes. The results from this study provide basic data for future research regarding the impacts of metal pollution on Apis mellifera and will be an important step towards a comprehensive risk assessment of the environmental stressors on honey bees.Chemosphere (2016), 164: 98-10

Anthropogenic influence on seasonal and spatial variation in bioelements and non-essential elements in honeybees and their hemolymph

Honeybee colony losses have been a focus of research in the last years, due to the importance of managed honeybee colonies for economy and ecology. Different unfavorable conditions from the outside environment have a strong impact on the hive health. The majority of losses occur mainly during winter and the exact reason is not completely understood. Only a small number of studies are dealing with content of bioelements, their function and influence on honeybee physiology. The aim of the present study was to determine seasonal and spatial variations in content of bioelements and non-essential elements, in hemolymph and whole body of honeybees originating from three regions with different degrees of urbanization and industrialization. Concentrations of 16 elements were compared: macroelements (Ca, K, Mg, Na), microelements (Cu, Fe, Mn, Zn) and non-essential elements (Al, Ba, Cd, Co, Cr, Ni, Pb, Sr) in samples collected from 3 different environments: Golija (rural region), Belgrade (urban region) and Zajača (industrial region). Content of bioelements and non-essential elements in honeybees was under noticeable influence of the surrounding environment, season and degree of honeybee activity. Hemolymph was proven to be helpful in differentiating air pollution from other sources of honeybee exposure. The results of our study demonstrated that bees can be successfully used as biomonitors since we have observed statistically significant differences among observed locations, but unless compared locations are exposed to excessively different pollution pressures, it is essential that all bees should be collected at the same season

Oxidative stress and the activity of antioxidative defense enzymes in overwintering honey bees

Over the past decades, the number of managed honey bee Apis mellifera L. (Hymenoptera: Apidae) colonies have been decreasing. The majority of losses occur during winter, suggesting that overwintering honey bees are more susceptible to adverse factors. We focused on the oxidative status of overwintering honey bees, particularly at the beginning (November) and end (March) of the wintering period. Colonies from three locations with different anthropogenic influences were selected: Belgrade, an urban zone, Zajača, an industrial zone, and Susek, a rural area. We measured levels of malondialdehyde (MDA), as a marker of lipid peroxidation, as well as the expression and activity of select antioxidative enzymes: superoxide dismutase (SOD), catalase (CAT) and glutathione S-transferase (GST). Our results show that enzyme activity and gene expression of antioxidative enzymes are influenced by both sample location and the time of sampling. The majority of analyzed genes had significantly reduced expression, at the end of the overwintering period when higher activities of antioxidative enzymes were also recorded. Among the analyzed parameters, SOD activity and gene expression of microsomal GST isoforms were more affected by local environmental conditions, suggesting the complex role of these enzymes in antioxidative defense and detoxification. The higher MDA levels observed at the end of overwintering for all three locations likely reflects accumulated oxidative damage which could be associated with the aging process, brood rearing and/or the onset flying activity

Environmental Effects on Superoxide Dismutase and Catalase Activity and Expression in Honey Bee

Understanding the cellular stress response in honey bees will significantly contribute to their conservation. The aim of this study was to analyze the response of the antioxidative enzymes superoxide dismutase and catalase in honey bees related to the presence of toxic metals in different habitats. Three locations were selected: (i) Tunovo on the mountain Golija, as control area, without industry and large human impact, (ii) Belgrade as urban area, and (iii) Zajaca, as mining and industrial zone. Our results showed that the concentrations of lead (Pb) in whole body of bees vary according to habitat, but there was very significant increase of Pb in bees from investigated industrial area. Bees from urban and industrial area had increased expression of both Sod1 and Cat genes, suggesting adaptation to increased oxidative stress. However, in spite increased gene expression, the enzyme activity of catalase was lower in bees from industrial area suggesting inhibitory effect of Pb on catalase