Ivana Tlak Gajger

Monitoring health status of Croatian honey bee colonies and possible reasons for winter losses

During recent years it appears that there have been increasing losses of honey bee colonies, and potential reasons are diseases, environmental factors such as pesticides and nutrition related stress among others. Honey bee diseases in particular pose considerable difficulties for beekeepers and the economy of a whole country. The intention of this study was to determine the total winter losses and the opinion of the beekeepers about the potential reasons for bee losses and the health conditions of their colonies.

Detection of Nosema ceranae in honey bees from Croatia

honey bee, distribution, Nosema apis, Nosema ceranae, Croatia Journal of Apicultural Research and Bee World 49(4): 340-341 (2010)

Zeolite clinoptilolite as a dietary supplement and remedy for honeybee (Apis mellifera L.) colonies

Control of the nosema disease poses a major challenge, and therefore, treatment of this serious para - sitic disease using natural preparations could be of great benefit. The aim of this study was to test the performance of zeolite clinoptilolite as a curative measure against honeybee colonies ( Apis mellifera L.) naturally infected by Nosema ceranae. The histopathological structure, and the content and distribution of mucosubstances and his - tochemical activity of aminopeptidase and non-specific esterase in the midgut mucosa of honeybees originating from colonies fed sugar syrup supplemented with zeolite minerals was studied. A decline in the number of spores in honeybees fed with zeolite clinoliptolite was observed on the first sampling day (Day 10; 53.25 15.15 million spores/bee), though a statistically lower number of spores in comparison to the control was confirmed on Day 20 (41.08 ± 9.4 million spores/bee), Day 30 (28.42 ± 7.79 million spores/bee) and Day 40 (24 ± 6.25 million spores/bee). The possibility of using natural zeolites as a dietary supplement for honeybee colonies as a preventative measure and for the reduction of the deleterious effects of nosemosis is discussed.

Essential and toxic element concentrations in monofloral honeys from southern Croatia

The concentrations of 24 elements in seven honey types (multifloral, heather, common heather, bearberry, sage, mandarin orange-blossom and honeydew) collected in southern Mediterranean regions of Croatia were determined using ICP-MS. Significant differences were found in the concentrations of Ag, As, Ba, Cu, Co, Fe, K, Mg, Mn, Mo, Na, Ni, Se, Sb, U and Th (p<0.05, all) among honeys. The highest element concentrations were determined in honeydew honeys, with the exception of multifloral (Ca, Cr, Mo, Se), common heather (Mg, Na), bearberry (Ba, Fe, Pb) and sage (Ag) honeys. Among the floral honeys, the highest concentrations were found in multifloral honey (Al, As, Be, Ca, Cr, Mn, Mo, Ni, Se, Th and U), common heather (Co, K, Mg, Na, V), sage (Ag, Cd, Cu), and bearberry (Ba, Fe, Pb, Sb, Zn). The results contribute to the evidence supporting the role of botanical origin on the elemental composition of honey.

Structural changes in the rat placenta during the last third of gestation discovered by stereology

Structural changes in the rat placenta during the last third of gestation were for the first time assessed by stereology. Fischer female rats were euthanized on the day 16 or day 19 of gestation, and 35 placentas were collected. Three randomly selected placentas from each group were stereologically analyzed for the absolute volume. The proportion of the glycogenic cells and the trophoblast giant cells (TGC) in the basal part of the placenta was calculated using volume density. The absolute volume of the rat placenta on the day 16 of gestation was determined as 0.0638 cm3. The labyrinth comprised 0.0274 cm3, the basal plate 0.0271 cm3 and the decidua 0.0093 cm3. On the day 19 of gestation, the absolute volume of the placenta was 0.1627 cm3, the labyrinth occupied 0.0922 cm3, the basal plate 0.0596 cm3 and the decidua 0.0109 cm3. The volume density of trophoblast giant cells was 0.174 cm0 on the day 16 and 0.107 cm0 on the day 19 of gestation. The glycogenic cells comprised 0.379 percentage of the basal plate on the day 16 and 0.236 on the day 19 of gestation. We conclude that the absolute volume of the whole placenta and the labyrinth has increased from day 16 to the day 19 of gestation. In contrast, the volume density of glycogenic cells and trophoblast giant cells was higher on the day 16 than on the day 19 of gestation, probably due to the intensive trophoblast invasion during that time.

Trueness and precision of the real-time RT-PCR method for quantifying the chronic bee paralysis virus genome in bee homogenates evaluated by a comparative inter-laboratory study

The Chronic bee paralysis virus (CBPV) is the aetiological agent of chronic bee paralysis, a contagious disease associated with nervous disorders in adult honeybees leading to massive mortalities in front of the hives. Some of the clinical signs frequently reported, such as trembling, may be confused with intoxication syndromes. Therefore, laboratory diagnosis using real-time PCR to quantify CBPV loads is used to confirm disease. Clinical signs of chronic paralysis are usually associated with viral loads higher than 108 copies of CBPV genome copies per bee (8 log10 CBPV/bee). This threshold is used by the European Union Reference Laboratory for Bee Health to diagnose the disease. In 2015, the accuracy of measurements of three CBPV loads (5, 8 and 9 log10 CBPV/bee) was assessed through an inter-laboratory study. Twenty-one participants, including 16 European National Reference Laboratories, received 13 homogenates of CBPV-infected bees adjusted to the three loads. Participants were requested to use the method usually employed for routine diagnosis. The quantitative results (n=270) were analysed according to international standards NF ISO 13528 (2015) and NF ISO 5725-2 (1994). The standard deviations of measurement reproducibility (SR) were 0.83, 1.06 and 1.16 at viral loads 5, 8 and 9 log10 CBPV/bee, respectively. The inter-laboratory confidence of viral quantification (+/- 1.96SR) at the diagnostic threshold (8 log10 CBPV/bee) was+/- 2.08 log10 CBPV/bee. These results highlight the need to take into account the confidence of measurements in epidemiological studies using results from different laboratories. Considering this confidence, viral loads over 6 log10 CBPV/bee may be considered to indicate probable cases of chronic paralysis.

Effects of short-term exposure to mobile phone radiofrequency (900 MHz) on the oxidative response and genotoxicity in honey bee larvae

Exposure of different animal species to radiofrequency electromagnetic fields (RF-EMF) could cause various biological effects such as oxidative stress, genotoxic effects and dysfunction of the immune system. However, there are a lack of results on oxidative stress response and genotoxicity in the honey bee (Apis mellifera) after exposure to RF-EMF. This study was performed to investigate the effects of exposure to RF-EMF on the activity of catalase, superoxide dismutase, glutathione S-transferase, lipid peroxidation level and DNA damage in honey bee larvae. Honey bee larvae were exposed to RF-EMF at 900 MHz and field levels of 10, 23, 41 and 120 V m−1 for 2 h. At a field level of 23 V m−1 the effect of 80% AM 1 kHz sinusoidal and 217 Hz modulation was investigated as well. Catalase activity and the lipid peroxidation level decreased significantly in the honey bee larvae exposed to the unmodulated field at 10 V m−1 compared to the control. Superoxide dismutase and glutathione S-transferase activity in the honey bee larvae exposed to unmodulated fields were not statistically different compared to the control. DNA damage increased significantly in honey bee larvae exposed to modulated (80% AM 1 kHz sinus) field at 23 V m−1 compared to the control and all other exposure groups. These results suggest that RF-EMF effects in honey bee larvae appeared only after exposure to a certain EMF conditions. The increase of the field level did not cause a linear dose-response in any of the measured parameters. Modulated RF-EMF produced more negative effects than the corresponding unmodulated field. Although honey bees in nature would not be exposed to such high field levels as used in our experiments, our results show the need for further intensive research in all stages of honey bee development.

Variations in lead, cadmium, arsenic, and mercury concentrations during honeybee wax processing using casting technology

Abstract Beeswax foundations are a necessary material in intensive modern beekeeping. Heavy metals can accumulate in these foundations for decades, as it is a common beekeeping practice to recycle wax. Beeswax samples were analysed using GFAAS for As, Cd, Pb, and Hg concentrations during the production of beeswax foundations using casting technology with a prolonged cooling and sedimentation phase. Significant differences were determined in the concentrations of As, Cd, Pb (p<0.01, all), and Hg (p<0.05) between the three levels of wax in a double-walled steel casting container and comb foundations (CF) during the processing stage. Concentrations (mg kg-1) of the examined metals in comb foundations as the final product ranged as follows: As 0.01-0.88; Cd 1.26-3.55; Pb 82.5-171, and Hg 0.29-1.46. All examined element concentrations demonstrated similar distribution and ratio in different layers, ranging from the lowest concentrations in layers from which wax material is used for comb foundations as the final product, to the highest concentrations in sedimented layer, which represents waste. The obtained results suggest that the described method could effectively eliminate a significant amount of heavy metals from the initial material used for the production of new beeswax foundations.

Reply to comment on "Structural changes in the rat placenta during the last third of gestation discovered by stereology"

Dear Editor, We are thankful to our colleagues for their interest in our paper and comments that they have provided. We could not agree more with the part of the letter regarding the method of sampling. On the basis of our 16-year-long experience in research on rat placentas, we underline sampling as one of the most important concepts in stereology, especially in the light of biological variability [1]. This variability is the reason why we approach sampling with the greatest care, as it is evident from all our published papers in the field of stereology. In addition, we consider it as one of the best methods for quantification of biological materials. Systematic random sampling, which combines both the unbiasedness of random sampling and the efficiency of a systematic sampling, was used in our research. It is based on the selection of the final sample systematically, while the first sample was selected randomly within the first sampling interval [2]. Sections were sampled at a ratio 1:10 (Section Sampling Fraction-SSF), with a random section (RS) selection from the first ten sections (in this case, the 3rd section) and then every 10th section after this initial selection. Preliminary measurements were performed for each of the experimental groups. In addition, due to its small size, it is possible to analyze the entire section of the rat placenta as a single field, which we did, as seen in Figure 2 [3]. However, this is not possible for human placenta, which, due to its larger size, cannot fit into a single section or single field for stereological analysis.