Beata Panasiuk

Cryopreservation of honeybee ( Apis melifera ) drone semen remains a significant challenge. Despite decades of research, maintaining sperm viability and functionality after thawing continues to limit the practical use of frozen semen. This study investigated the effect of post-thaw dilution of cryopreserved drone semen with Bee Semen Solution (BSS) on semen viability and queen reproductive performance. Queen bees were instrumentally inseminated with fresh or cryopreserved semen, either undiluted or diluted with BSS after thawing. The impact of these treatments on the number and quality of spermatozoa in the spermatheca and on brood pattern was assessed. Cryopreservation markedly affected sperm quality and the reproductive performance of inseminated queens, resulting in lower sperm viability and reduced fertility compared with fresh semen. However, post-thaw dilution of semen with BSS tended to improve sperm parameters in the spermatheca and queen fertility. The results indicate that post-thaw handling of semen, including the use of appropriate diluents, may play an important role in improving the outcomes of insemination with dialysis method cryopreserved semen. We also emphasize that multiple insemination should be considered as a method of improving spermatozoa number in spermahteca and, consequently, queen fertility.

In the fight against the Varroa destructor mite, selective breeding of honey bee (Apis mellifera L.) populations that are resistant to the parasitic mite stands as a sustainable solution. Selection initiatives indicate that using the suppressed mite reproduction (SMR) trait as a selection criterion is a suitable tool to breed such resistant bee populations. We conducted a large European experiment to evaluate the SMR trait in different populations of honey bees spread over 13 different countries, and representing different honey bee genotypes with their local mite parasites. The first goal was to standardize and validate the SMR evaluation method, and then to compare the SMR trait between the different populations. Simulation results indicate that it is necessary to examine at least 35 single-infested cells to reliably estimate the SMR score of any given colony. Several colonies from our dataset display high SMR scores indicating that this trait is present within the European honey bee populations. The trait is highly variable between colonies and some countries, but no major differences could be identified between countries for a given genotype, or between genotypes in different countries. This study shows the potential to increase selective breeding efforts of V. destructor resistant populations.

The identification of honey bee ( Apis mellifera ) subspecies is often based on the measurements of workers’ fore-wings. The interpretation of the measurements can be difficult because the phenotype of workers is affected by both genetic and environmental factors. Moreover, it is not clear how the phenotype is affected by maternal inheritance. We have used the methodology of geometric morphometrics to verify if hybrids of honey bee subspecies and their backcrosses are more similar to either the father or mother colony. The comparison was based on fore-wing venation of three honey bee subspecies: A. m. carnica, A. m. caucasica, A. m. mellifera . First generation hybrids and backcrosses of those subspecies were obtained through instrumental insemination. Workers of the hybrids were compared with their parental colonies. The shape of wing venation was more similar to the maternal than to parental colony. This phenomenon was particularly visible in first generation of hybrids but it was also present in backcrosses. There were also symptoms of genetic dominance of some subspecies but this effect interacted with maternal inheritance and was difficult to interpret.

During natural mating honeybee queens can get lost due to drifting, predators or other cases. In this work, the balling of queens returning from flights by worker bees originating from the same colony was observed. Three subspecies of bees Carniolan, Caucasian and European Black Bee were tested. Research was conducted in both spring and summer, but in the former in newly created colonies, while in the latter in new and earlier used ones. Generally 15.2% of queens were balled and in total 30.2% of queens were lost during mating flights. 269 queens performed 785 mating flights, and 5.2% of those finished with balling. Three times more queens were balled when returning from mating flight rather than orientation flight. Subspecies matches or mismatches of queens and workers in nucleuses did not significantly affect the balling or its frequency. Additionally, no bee subspecies characterized stronger tendencies to ball a queen. Worker bees from newly created nucleuses treated queens similarly to the ones in nucleuses earlier used. However, significantly more queens had been balled during the spring in comparison to summer. There were days with higher balling of queens. During some days the weather was very unstable and unpredictable with such anomalies as heat waves, thunderstorms or sudden drops in insolation. Most of the queens were balled at the entrance while returning from flight and only a few inside the hive. In the research, clear causes of balling were not found, but some factors can be excluded.

The aim of this study was to examine whether the addition of a diluent for honey bee sperm, directly before insemination, affects the quality and vitality of queens and their brood, in comparison to queens inseminated traditionally, with fresh semen. Honey bee queens were inseminated with diluted and homogenized semen collected from a few dozen drones, in order to increase the diversity of their offspring. Queens and drones (Apis mellifera carnica) were mated within subspecies, however, only three selected lines were used. The average number of spermatozoa filling the spermatheca of queens, inseminated with semen without dilution, freshly collected from drones, from a single colony (SCS) was not significantly higher, compared to queens inseminated with semen diluted and mixed (MCS); amounting to 5.1 and 4.4 million sperm, respectively. Queens from both groups started laying eggs at a similar time after insemination, and their average life expectancy in colonies, was 20.5 and 22.6 months, respectively. The most common reason for queen losses was supersedure, and the reasons for these losses were not significantly different between groups. Brood survival rates, in both groups, did not differ significantly.

Honey bee queens were inseminated with diluted, homogenised semen collected from a few dozen drones. This procedure was carried out to increase the diversity of the queens’ offspring, which is in comparison to the offspring of queens inseminated with semen from only a few drones coming from one colony. Queens and drones were mated within carniolan bee (Apis mellifera carnica) subspecies, but 3 selected lines were used. Queens were reared from one line and drones from the same line, and two additional lines differing in hygienic behaviour wherein in one of them that trait was strongly evident. The aim of this study was to examine whether the level of enhanced genetic variability in colonies and simultaneously the participation of hygienic bees, would increase the performance of hygienic behaviour. Overall hygienic behaviour of colonies with a lower and greater genetic variability did not differ significantly and amounted to 52.1 and 47.0%, respectively. Colonies within the lower variability group, in which drones from line selected in hygienic behaviour performance were used for inseminating queens, had a significantly greater percent of cleaned pupae than other colonies (63.2%). Hygienic behaviour in other colonies was more dependent on the gene quotas of hygienic bees in the colonies rather than on the level of polyandry.

The aim of the study was to determine the effect of honey bee worker diversity within the colony on: development, honey productivity, and wintering. Two different levels of diversity within the colony were tested. The appropriate levels of diversity within the colony were obtained by selecting drones for inseminating the queens. Lower genetic diversity was obtained in the colonies headed by a queen inseminated with semen collected from drones originating from a single colony. Higher genetic diversity was obtained in the colonies with queens inseminated with semen from drones of 30 different colonies. Colonies with a higher genetic variation of workers in the colonies had greater levels of functional characteristics. However, apart from the number of dead bees in winter, the genetic diversity level of the workers on the colony development and honey production, did not have a significant influence. There was an averaging effect observed concerning that male component in the colonies with a higher genetic variation of workers - on honey yield, when compared to the non-additive effect of the best drones.

Winter honey bee losses were evaluated during the two overwintering periods of 2009/2010 and 2010/2011. The research included dead bee workers that fell on the hive bottom board (debris) and the ones that flew out of the hive. Differences were observed in the number of bees fallen as debris between the two periods, whereas the number of bees flying out was similar in both years. No differences were found between the numbers of dead bees in strong and weak colonies. The percentage of bees flying out of the colony increased in the presence of Nosema spores, Varroa infestation, increased average air temperature, and insolation during the day. In addition, both the presence of Nosema and insolation during the day had an impact on the number of bees that died and fell on the hive board.

The susceptibility of bee larvae to Ascosphaera apis infestation and the hygienic behaviour of worker bees in relation to A. apis infected and freeze-killed brood were evaluated in three races of bees: Apis mellifera carnica, Apis mellifera caucasica, and Apis mellifera mellifera. Experimental bee colonies were evaluated in field conditions during the three beekeeping seasons. The lowest percentage of infected larvae was observed in car GR1 and mel A colonies (8.5% and 15%, respectively) and the highest in car Mr and cau P colonies (21% and 24.3%, respectively). Bees in the car GR1 and mel A colonies removed mummified brood in a shorter period of time (6.5 and 7.1 days on average, respectively) than car Mr and cau P colonies (above 8 days). Bees in the mel A and car GR1 colonies cleaned significantly more cells with freeze-killed brood within 24 and 48 hours (above 70% and 80% on average, respectively) than car Mr and cau P colonies (on average 10 - 20% lower cleaning rate). A low correlation coefficient was found for the susceptibility of larvae to A. apis infection and hygienic behaviour.