Research Article
Bożena Chuda-Mickiewicz 
, Jerzy Samborski
Department of Zoology and Apiculture, West Pomeranian University of Technology in Szczecin, 20 Doktora Judyma St., 71-466 Szczecin
Abstract. The quality of queens reared in colonies with restricted (RP) and not restricted pollen supply (NRP) was evaluated during foraging on false acacia and small-leaved and broad-leaved lime. It was shown that the body weight of queens in both groups immediately after emergence and on the day of instrumental insemination did not differ significantly (221.8 ± 15 and 224.3 ± 15 and 170.9 ± 15 and 177.5 ± 14 mg, respectively). The waiting time to start oviposition was similar: 4.7 ± 1.8 days in RP group and 5.7 ± 2.8 days in NRP group, not being significantly different. There were no significant differences in the diameter and volume of spermatheca and the number of ovarioles in the right ovary between the queens of the tested groups.
Keywords: Apis mellifera L., queen, pollen, instrumental insemination
The honey bee food is nectar, honeydew and pollen [Haydak 1970Haydak, M.H. (1970). Honey bee nutrition. Annual Rev. Entomology, 15, 143–156. https://doi.org/10.1146/annurev.en.15.010170.001043]. Nectar and honeydew contain mainly carbohydrates. Pollen, on the other hand, is the main source of proteins, fats as well as minerals and vitamins [Roulston and Cane 2000Roulston, T.H., Cane, J.H. (2000). Pollen nutritional content and digestibility for animals. Plant Syst. Evol., 222, 187–209. https://doi.org/10.1007/BF00984102]. Freshly harvested pollen as well as that stored in comb cells (in the form of beebread) is the food mainly for young nurse bees [Crailsheim et al. 1992Crailsheim, K., Schneider, L.H.W., Hrassnigg, N., Bühlmann, G., Brosch, U., Gmeinbauer, R., Schöffmann, B. (1992). Pollen consumption and utilization in worker honeybees (Apis mellifera carnica):dependence on individual age and function. J. Insect Physiol., 38, 409–419. https://doi.org/10.1016/0022-1910(92)90117-V].
Pollen is the only source of proteins that guarantee the proper development and functioning of many honeybee organs [Haydak 1970Haydak, M.H. (1970). Honey bee nutrition. Annual Rev. Entomology, 15, 143–156. https://doi.org/10.1146/annurev.en.15.010170.001043]. The amount and quality of pollen consumed by nurse bees is the basis for the proper development of the hypopharyngeal glands located in the head [Pernal and Currie 2000Pernal, S., Currie, R.W. (2000). Pollen quality of fresh and 1-year-old single pollen diets for worker honey bees (Apis mellifera L.). Apidologie, 31, 387–409. https://doi.org/10.1051/apido:2000130, Renzi et al. 2016Renzi, M.T., Rodríguez-Gasol, N., Medrzycki, P., Porrini, N., Martini, A., Burgio, G., Maini, S., Golastra, F. (2016). Combined effect of pollen quality and thiamethoxam on hypopharyngeal gland development and protein content in Apis mellifera. Apidologie, 47, 779–788. https://doi.org/10.1007/s13592-016-0435-9] that produce royal jelly. High-protein food used to feed larvae developing into worker bees and drones up to day 3 of life or until the end of the stage of larvae developing in queens (reared queens). It is also the food of adult queens throughout their entire live [Crailsheim 1992Crailsheim, K. (1992). The flow of jelly within a honeybee colony. J. Comp. Physiol. B, 162 (8), 681–689. https://doi.org/10.1007/BF00301617, Fujita et al. 2013Fujita, T., Kozuka-Hata, H., Ao-Kondo, H., Kunieda, T., Oyama, M., Kubo, T. (2013). Proteomic analysis of the royal jelly and characterization of the functions of its derivation glands in the honeybee. J. Proteome Res., 12, 404−411. https://doi.org/10.1021/pr300700e]. The protein diet is mainly consumed by 1–8 day old bees, with the highest consumption on day 3. The acini of hypopharyngeal glands attained the age of 5–8 days and then began to shrink afterwards [Deseyn and Billen 2005Deseyn, J., Billen, J. (2005). Age-dependent morphology and ultrastructure of the hypopharyngeal gland of Apis mellifera workers (Hymenoptera, Apidae). Apidologie, 36, 49–57. https://doi.org/10.1051/apido:2004068, Omar et al. 2017Omar, E., Abd-Ella, A.A. Khodairy, M.M., Mosbeckhofer, R., Crailsheim, K., Brodschneider, R. (2017). Influence of different pollen diets on the development of htpopharyngeal glands and size of acid gland sacs in caged honey bees (Apis mellifera). Apidologie, 48, 425–436. https://doi.org/10.1007/s13592-016-0487-x].
The decreased availability of pollen as well as its low quality contributes to the development of smaller hypopharyngeal glands [DeGrandi-Hoffman et al. 2010DeGrandi-Hoffman, G., Chen, Y.P., Huang, E., Huang, M.H. (2010). The effect of diet on protein concentration, hypopharyngeal gland development and virus load in worker honey bees (Apis mellifera L.). J. Insect Physiol., 56(9), 1184–1191. https://doi.org/10.1016/j.jinsphys.2010.03.017, Renzi et al.2016]. In the rainy season, and during the period of no pollen supply, the frequency of visiting young larvae (1–3 days old) by nurse bees is smaller and correlated with the amount of pollen in the brood chamber and the number of non-capped larvae [Schmickl and Crailsheim 2002Schmickl, T., Crailsheim, K. (2002). How honeybee (Apis mellifera L.) change their broodcare behavior in response to non-foraging condition and poor pollen conditions. Behav. Ecol. Sociobiol., 51, 415–425. https://doi.org/10.1007/s00265-002-0457-3]. Bad pollen supply also led to cannibalism of larvae less than 3 days old and to shortening of the time until larvae are sealed [Schmickl and Crailsheim 2001Schmickl, T., Crailsheim, K. (2001). Cannibalism and early capping: strategy of honeybee colonies in times of experimental pollen shortages. J. Comp. Physiol., A 187, 541–547. https://doi.org/10.1007/s003590100226]. Bees rear less brood [DeGrandi-Hoffman et al. 2008DeGrandi-Hoffman, G., Wardell, G., Ahumada-Segura, F., Rinderer, T., Danka, R., Pettis J. (2008). Comparisons of pollen substitute diets for honey bees: consumption rates by colonies and effects on brood and adult populations. J. Apic. Res., 47(4), 265–270. https://doi.org/10.3896/IBRA.1.47.4.06] and the life span of bees is shorter [Di Pasquale et al. 2013Di Pasquale, G., Salignon, M., Le Conte, Y., Belzunces, L.P., Decourtye, A., Kretzschmar, A., Suhail, S., Brunet, J., Alaux, C. (2013). Influence of pollen nutrition on honey bee health: do pollen quality and diversity matter?. PLoS ONE, 8(8), e72016. https://doi.org/10.1371/journal.pone.0072016].
The aim of our study was to evaluate and compare the robustness and onset of oviposition in queens reared in colonies with restricted and not restricted pollen supply.
The study was conducted at the Department of Zoology and Apiculture, West Pomeranian University of Technology in Szczecin in full beekeeping season (June-July 2015) during foraging on false acacia and small-leaved and broad-leaved lime. The research material consisted of sister queens of Apis mellifera carnica being reared from one-day larvae [Ruttner and Ruttner 1983Ruttner, H., Ruttner, F. (1983). Reliable rearing methods [In:] Queen rearing -- biological basis and technical instruction. Apimondia Publishing House, Bucharest. Google Scholar, Büchler et al. 2013Büchler, R., Andonov, S., Bienefeld, K., Costa, C., Hatjina, F., Kezic, N., Kryger, P., Spivak, M., Uzunov, A., Wilde, J. (2013). Standard methods for rearing and selection Apis mellifera queens. J. Apic. Res., 52(1), 1–9. https://doi.org/10.3896/IBRA.1.52.1.07].
Queens were reared in two queenless colonies colonising Wielkopolski hives (360×260 mm frame) with a bottom pollen trap with 5.0 mm diameter hols, collecting about 29% pollen loads [Bobrzecki and Wilde 1990Bobrzecki, J., Wilde, J. (1990). Collection and utilisation of pollen loads [Pozyskiwanie i zagospodarowanie obnóży pyłkowych]. PWRiL Poznań [in Polish]. Google Scholar]. In one colony, 10 days before the beginning of rearing, pollen load trapping was started. Queen cells from the colony with restricted (RP) and not restricted pollen supply (NRP) after capping were placed in an incubator at 34.5°C. Staring from day 15 of preimaginal development, the emergence of queens was monitored every six hours. Immediately after emergence, queens were weighed (on a RADWAG WXD 200/2000 laboratory balance for weighing the moving animals) with an accuracy to 1 mg and introduced into plastic mailing cages with 8–10 bees and with candy. On the second day of life, queens were introduced into mini-plus mating nuclei (with 215×163 mm frame), with bees occupaiting 6 combs, including two with capped brood. The nuclei entrances were secured with a queen excluder.
At the age of seven days, queens – after weighing (on a RADWAG WXD 200/2000 laboratory balance, with accuracy to 1 mg) – were instrumentally inseminated once with 8 µl of semen [Woyke 1960Woyke, J. (1960). Natural and instrumental insemination of queen bees [Naturalne i sztuczne unasienianie matek pszczelich]. Pszczeln. Zesz. Nauk., 4(3-4), 183–275 [in Polish]. Google Scholar, Cobey et al. 2013Cobey, S.W., Tarpy, D.R., Woyke, J. (2013). Standard methods for instrumental insemination of Apis mellifera queens. J. Apic. Res., 52(4). https://doi.org/103896/IBRA.1.524.09] collected from drones of the same subspecies aged 14–16 days [Woyke and Jasiński 1978Woyke, J., Jasiński, Z. (1978). Influence of age of drones on the results of instrumental insemination of honeybee queens. Apidologie, 9(3), 203–212. https://doi.org/10.1051/apido:19780304]. Two days after instrumental insemination, queens were anaesthetised with CO2 for 3 minutes [Woyke et al. 2001Woyke, J., Fliszkiewicz, Z., Jasiński, Z. (2001). Prevention of natural mating of instrumentally inseminated queen honeybees by proper method of instrumental insemination. J. Apic. Sci., 45, 101–114. Google Scholar]. The onset of oviposition by queens was monitored by examining the mating nuclei every two days, until day 14 after instrumental insemination.
In randomly selected egg lying queens from both groups, after anaesthetising, the reproductive system was removed [Dade 2009Dade, H.A., (2009). Dissection of the queen [In:] Anatomy and dissection of the honeybee. International Bee Research Association, 137–140. Google Scholar]. The diameter of spermatheca was determined by a modified method of Woyke [1971]Woyke, J. (1971). Correlation between the age at which honeybee brood was grafted, characteristics of the resultant queens, and result of insemination. J. Apic. Res., 10(1), 45–55. https://doi.org/10.1080/00218839.1971.11099669. The removed spermathecae were transferred onto microscope glass slides. After removing the trachea, the photographs of spermathecae were taken using a Zeiss Primo Star microscope with a digital camera (at 20× magnification) and their diameter were measured by means of ScopePhoto software. The number of ovarioles in the right ovary was counted [Carreck et al. 2013Carreck, N.L., Andree, M., Brent, C.S, Cox-Fster, D., Dade, H.A., Ellis, J.D., Hatjina, F., Englesdory, D. (2013). Standard methods for Apis mellifera anatomy and dissection. J. Apic. Res, 52(4), 1–39. https://doi.org/10.3896/IBRA.1.52.4.03]. The removed ovaries were submerged in 95% alcohol for 15 minutes, transferred onto microscope glass slides and cut at half their length [Woyke 1971Woyke, J. (1971). Correlation between the age at which honeybee brood was grafted, characteristics of the resultant queens, and result of insemination. J. Apic. Res., 10(1), 45–55. https://doi.org/10.1080/00218839.1971.11099669]. In the cut off thicker, distal part, ovarioles were counted under a microscope, at 25× magnification.
In total, the body weight and the onset of oviposition were evaluated in 61 queens, including 32 in RP group and 29 in NRP group. The spermathecal diameter and the ovariole number in the right ovary were determined in 10 queens in each group.
The figures obtained were analysed statistically using Statistica PL v.12 computer software. Differences between the body weight of queens, diameter and volume of their spermathecae, and the number of ovarioles were evaluated with the Student’s t-test. Differences in the waiting time to start oviposition by queens was analysed with a χ2 test. The coefficients of correlation between the physical traits being examined were calculated. The number of queens starting oviposition was analysed with the G-test with Williams’s correction [Sokal and Rohlf 1981Sokal, R.R., Rohlf, F.J. (1981). Biometry. W H Freeman and Company, New York, USA. Google Scholar].
Mean body weight (±SD) of queens immediately after emergence (on day 1 of life) in NRP group was only by 2.5 mg greater than in RP group (Table 1). Similarly, on the day of instrumental insemination, the queens from NRP group had a slightly greater body weight, by 6.6 mg on average, than the queens from RP group (Table 1). The observed differences in queen body weight between the groups both after emergence and on the day of instrumental insemination were not significant (t = –0.632, P = 0.529 and t = 1.789, P = 0.0787). In the period of maturation from emergence to instrumental insemination, the body weight of queens in RP group and NRP group decreased significantly (t = 13.554, P = 0.000 and 12.229, P = 0.000), by 22.9 and 20.9%, respectively (Table 1).
Out of 32 instrumentally inseminated queens in RP group and 29 in NRP group, until day 14 after instrumental insemination, a similar, not significantly different number of queens started oviposition (G-test: Gadj = 0.001, padj = 0.974), i.e. 93.7 and 93.2%, respectively. In RP group, two queens were found missing on days 6 and 8 after instrumental insemination, while in NRP group one queen was missing on day 10 and another one did not start oviposition. Her dissection did not show any residual semen in the lateral oviducts, while the spermatheca was filled with sperm.
The waiting time to start oviposition by queens in both groups was similar, not being significantly different (χ2 = 0.335, P = 0.563). The modal value for the number of days from instrumental insemination to the onset of oviposition in RP group was by 1 day smaller than in NRP group, in which it was 5 days. The evaluation of the reproductive system of queens did not show significant differences in the spermathecal diameter (t = –0.935, P = 0.362), its volume (t = –0.973, P = 0.342), and the ovariole number in the right ovary (t = 0.083, P = 0.935) (Table 1).
Table
1. Average values (±SD)
traits investigated in queens from groups with restricted (RP)
and not restricted pollen supply (NRP). |
||||
|
N |
Restricted pollen supply |
N |
Not restricted pollen supply |
Body weight immediately after emergence, mg |
32 |
221.8 ±15.1 aA* |
29 |
224.3 ±15.3 aA |
Body weight prior to instrumental insemination, mg |
32 |
170.9 ±15.0 aB |
29 |
177.5 ±13.8 aB |
Number of days from instrumental insemination to starting
oviposition |
30 |
4.7 ±1.8 a |
27 |
5.7 ±2.8 a |
Spermathecal diameter, mm |
10 |
1.11 ±0.02 a |
10 |
1.12 ±0.03a |
Spermathecal volume, µl |
10 |
0.714 ±0.04 a |
10 |
0.737 ±0.06 a |
Ovariole number in the right ovary |
10 |
127.2 ±16.2 a |
10 |
126.7 ±10.1 a |
* values marked with different lowercase letters in rows and
uppercase ones in columns differ significantly at p = 0.05. |
||||
No significant differences between the parameters evaluated in queens being reared in colonies with restricted (RP) and not restricted pollen supply (NRP) allowed for a joint analysis of correlations between them. The correlation coefficients calculated between the body weight of queens immediately after emergence as well as on the day of instrumental insemination and the onset of oviposition by queens, r = 0.100 and r = 0.108, were not significant (t = 745, P = 0.459 and t = 0.803, P = 0.425). Similarly, no correlation was found between the body weight of queens immediately after emergence and the ovariole number in the right ovary, r = –0.267 (t = –1.172, P = 0.256), and the spermathecal diameter and volume – r = 0.261 (t = 1.150, P = 0.296) and r = 0.262 (t = 1.150, P = 0.265), as well as the ovariole number in the right ovary and the spermathecal diameter and volume, r = –0.03 (t = –0.398, P = 0.696) and r = –0.107 (t = –0.461, P = 0.651).
While undertaking the study, it was presumed that a restricted 10-day pollen supply o to a colony before and during the queen rearing (with pollen trapping) would affect the quality of queens being reared. The analysis of the findings did not confirm our assumptions. The body weight of queens immediately after emergence and on day 7 of life, just before instrumental insemination in RP group and NRP group did not differ significantly. The emerging queens in both groups were heavier on average by 28 to 57 mg when compared to the studies by Bieńkowska et al. [2009]Bieńkowska, M., Panasiuk, B., Gerula, D., Węgrzynowicz, P. (2009). Weight of honeybee queens and its effect on the quality of instrumentally inseminated queens. Proceedings of the 4th International Apicultural Congress, Montpellier, France, 15–20 September 2009, p. 135. Google Scholar and Alqarni et al. [2013]Alqarni, A.S., Balhareth, H.M., Owayss, A.A. (2013). Queen morphometric and reproductive characters of Apis mellifera jemenitica, a native honey bee to Saudi Arabia. Bull. Insec., 66(2), 239–244, Google Scholar, respectively. The body weight of Carniolan queens immediately after emergence, similar to our results, i.e. over 200 mg, was found by Chuda-Mickiewicz and Samborski [2015]Chuda-Mickiewicz, B., Samborski, J. (2015). The quality of honey bee queens from queen cells incubated at different temperatures. Acta Sci. Pol. Zootechnica, 14(4), 3–10. Google Scholar and Skowronek et al. [2004]Skowronek, W., Bieńkowska, M., Kruk, C. (2004). Changes in body weight of honeybee queens during their maturation. J. Apic. Sci., 48(2), 61–68. Google Scholar. A significant queen body weight loss in RP and NRP groups, during maturation, before instrumental insemination, on day 7 of life, was on average lower by 4.2 to 8% when compared to the queens emerging from queen cells incubated at 34.5 and 32°C, respectively, and kept until instrumental insemination in mating nuclei in the study by Chuda-Mickiewicz and Samborski [2015]Chuda-Mickiewicz, B., Samborski, J. (2015). The quality of honey bee queens from queen cells incubated at different temperatures. Acta Sci. Pol. Zootechnica, 14(4), 3–10. Google Scholar. Similarly, Bienkowska et al. [2009] reported a smaller body weight loss, by 4.7%, in queens in the same period, being kept in cages with 25 bees. However, a larger loss of body weight, by 4.2%, from emergence to instrumental insemination, when compared to the present study and those of earlier cited authors, was stated by Skowronek et al. [2004]Skowronek, W., Bieńkowska, M., Kruk, C. (2004). Changes in body weight of honeybee queens during their maturation. J. Apic. Sci., 48(2), 61–68. Google Scholar, who kept queens in nuclei. The body weight of queens on the day of instrumental insemination in both groups was within the range given in the literature. i.e. 160 to 191 mg [Skowronek et al. 2004Skowronek, W., Bieńkowska, M., Kruk, C. (2004). Changes in body weight of honeybee queens during their maturation. J. Apic. Sci., 48(2), 61–68. Google Scholar, Bieńkowska et al. 2009Bieńkowska, M., Panasiuk, B., Gerula, D., Węgrzynowicz, P. (2009). Weight of honeybee queens and its effect on the quality of instrumentally inseminated queens. Proceedings of the 4th International Apicultural Congress, Montpellier, France, 15–20 September 2009, p. 135. Google Scholar, Chuda-Mickiewicz et al. 2012Chuda-Mickiewicz, B., Czekońska, K., Samborski, J., Rostecki, P. (2012). Success rates for instrumental insemination of carbon dioxide and nitrogen anaesthetised honey bee (Apis mellifera) queens. J. Apic. Res., 51(1), 74–77. https://doi.org/10.3896/IBRA.1.51.1.09, Heljanka et al. 2014, Chuda-Mickiewicz and Samborski 2015Chuda-Mickiewicz, B., Samborski, J. (2015). The quality of honey bee queens from queen cells incubated at different temperatures. Acta Sci. Pol. Zootechnica, 14(4), 3–10. Google Scholar].
The percentage of queens starting oviposition in our experiment was higher by 8–16% when compared to that being found by Mackensen [1947]Mackensen, O. (1947). Effect of carbon dioxide on initial oviposition of artificial inseminated and virgin queen bees. J. Econ. Entomol., 40, 344–349. https://doi.org/10.1093/jee/40.3.344, Kanfalung and Peng [1982]Kanfalung, O., Peng, Y.S. (1982). Effect of insemination on the initiation of oviposition in the queen honey bee. J. Apic. Res., 21, 3–6. https://doi.org/10.1080/00218839.1982.11100508, Czekońska and Chuda-Mickiewicz [2007]Czekońska, K., Chuda-Mickiewicz, B. (2007). Effect of induces defecation on the effectiveness of insemination in honey bee (Apis mellifera) queens. J. Apic. Res., 46(4), 242–244. https://doi.org/10.1080/00218839.2007.11101402, and Chuda-Mickiewicz et al. [Chuda-Mickiewicz et al. [2009]Chuda-Mickiewicz, B., Prabucki, J., Samborski, J., Rostecki, P. (2009). The role of phytohormones ininstrumental insemination of queen bees. J. Apic. Sci., 53(2), 91–96. Google Scholar, Chuda-Mickiewicz et al. [2012]Chuda-Mickiewicz, B., Czekońska, K., Samborski, J., Rostecki, P. (2012). Success rates for instrumental insemination of carbon dioxide and nitrogen anaesthetised honey bee (Apis mellifera) queens. J. Apic. Res., 51(1), 74–77. https://doi.org/10.3896/IBRA.1.51.1.09] for the queens inseminated on day 7 of life and anaesthetised with CO2 for 3 minutes two days before or after instrumental insemination. The effectiveness of instrumental insemination, similar to that in our study, was obtained by Bieńkowska et al. [2012]Bieńkowska, M., Panasiuk, B., Gerula, D., Węgrzynowicz, P. (2012). Effect of different carbon dioxide gas concentrations used during the insemination of honey bee queens in staring oviposition. J. Apic. Sci., 56(1), 125–135. https://doi.org/10.2478/v10289-012-0017-7, 92.6%, Chuda-Mickiewicz and Prabucki [2000]Chuda-Mickiewicz, B., Prabucki, J. (2000). Rifampicin, a new antibiotic in insemination of queen bees, Pszczel. Zesz. Nauk., 42(2), 87–94 Google Scholar and Chuda-Mickiewicz and Samborski [2015]Chuda-Mickiewicz, B., Samborski, J. (2015). The quality of honey bee queens from queen cells incubated at different temperatures. Acta Sci. Pol. Zootechnica, 14(4), 3–10. Google Scholar, 93.48 and 95%, respectively, and Gerula et al. [2011]Gerula, D., Bieńkowska, M., Panasiuk, B. (2011). Instrumental insemination of honey bee queens during flight activity predisposition period 1. Onset of oviposition. J. Apic. Sci., 55(2), 53–66. https://doi.org/10.2478/v10289-012-0016-8, 97.4%. A similar percentage of queens who started oviposition received Moritz and Kühnert [1984]Moritz, R.F.A., Kühnert, M. (1984). Seasonal effects on artificial insemination of honeybee queens (Apis mellifera L.). Apidologie, 15(2), 223–231. https://doi.org/10.1051/apido:19840210, i.e. 92.3, who used a longer 10-minute CO2 anaesthesia two days after instrumental insemination. According to Otten et al. [1998]Otten, C., Otto, A., Renner, R . (1998). Artificial insemination: Methodological influence on the results. Apidologie, 29, 476. Google Scholar, the onset of oviposition by inseminated queens is to a large extent determined by the number of bees in a mating nuclei. In strong colonies with 1200 worker bees, 95% of queens started oviposition, while in weak ones with 200 worker bees only 70% of queens. Otten et al. [1998]Otten, C., Otto, A., Renner, R . (1998). Artificial insemination: Methodological influence on the results. Apidologie, 29, 476. Google Scholar also demonstrated that it depends on the age at which the queens are inseminated; from those being inseminated on day 10 of life, 95% started oviposition, while those inseminated on days 7 and 13 were fewer, 82 and 80%, respectively. Similar results were obtained by Chuda-Mickiewicz and Prabucki [1993]Chuda-Mickiewicz, B., Prabucki, J. (1993). Undertaking the oviposition by honey bee queens kept in cages with non-free flaying bees [Podejmowanie czerwienia przez inseminowane matki pszczele przetrzymywane w skrzynkach w asyście swobodnie oblatujących się pszczół]. Pszczel. Zesz. Nauk., 37, 23–31 [in Polish]. Google Scholar who inseminated the queens twice with 4 μl of semen. Out of the queens inseminated at the age of 11 days, oviposition started 100%, of those inseminated at the age of 8 days 87.3%, and of those inseminated at the age of 6 days 50% to day 14 from the first insemination procedure. Mean waiting time to start oviposition in both RP group (4.7 days) and NRP group (5.7 days) was much shorter when compared to the study by Chuda-Mickiewicz et al. [Chuda-Mickiewicz et al. 2009Chuda-Mickiewicz, B., Prabucki, J., Samborski, J., Rostecki, P. (2009). The role of phytohormones ininstrumental insemination of queen bees. J. Apic. Sci., 53(2), 91–96. Google Scholar, Chuda-Mickiewicz et al. 2012Chuda-Mickiewicz, B., Czekońska, K., Samborski, J., Rostecki, P. (2012). Success rates for instrumental insemination of carbon dioxide and nitrogen anaesthetised honey bee (Apis mellifera) queens. J. Apic. Res., 51(1), 74–77. https://doi.org/10.3896/IBRA.1.51.1.09], 8.1 days, Gerula et al. [2011]Gerula, D., Bieńkowska, M., Panasiuk, B. (2011). Instrumental insemination of honey bee queens during flight activity predisposition period 1. Onset of oviposition. J. Apic. Sci., 55(2), 53–66. https://doi.org/10.2478/v10289-012-0016-8, 8.3 days, Czekońska and Chuda-Mickiewicz [2007]Czekońska, K., Chuda-Mickiewicz, B. (2007). Effect of induces defecation on the effectiveness of insemination in honey bee (Apis mellifera) queens. J. Apic. Res., 46(4), 242–244. https://doi.org/10.1080/00218839.2007.11101402, 9.9 days, and Skowronek et al. [2002]Skowronek, W., Kruk, C., Kłopot, J. (2002). Factors affecting oviposition of artificially inseminated honeybee queens. J. Apic. Res., 46(2), 85–95. Google Scholar, 10 days for the queens – as in the present study – being inseminated on day 7 of life and anaesthetised with CO2 for 3 minutes two days after instrumental insemination. Similarly, queens inseminated with 2×4 μl of semen started oviposition later, according to Chuda-Mickiewicz and Prabucki [Chuda-Mickiewicz and Prabucki 1993Chuda-Mickiewicz, B., Prabucki, J. (1993). Undertaking the oviposition by honey bee queens kept in cages with non-free flaying bees [Podejmowanie czerwienia przez inseminowane matki pszczele przetrzymywane w skrzynkach w asyście swobodnie oblatujących się pszczół]. Pszczel. Zesz. Nauk., 37, 23–31 [in Polish]. Google Scholar, Chuda-Mickiewicz and Prabucki 2000Chuda-Mickiewicz, B., Prabucki, J. (2000). Rifampicin, a new antibiotic in insemination of queen bees, Pszczel. Zesz. Nauk., 42(2), 87–94 Google Scholar], i.e. after 10.4 and 8.1 days, respectively, and Woyke et al. [2001]Woyke, J., Fliszkiewicz, Z., Jasiński, Z. (2001). Prevention of natural mating of instrumentally inseminated queen honeybees by proper method of instrumental insemination. J. Apic. Sci., 45, 101–114. Google Scholar – after 6–7 days. The faster oviposition of queens in our experiment was more likely to have been affected by the presence of capped brood and emerging bees in a the nucleus colony [Skowronek et al. 2002Skowronek, W., Kruk, C., Kłopot, J. (2002). Factors affecting oviposition of artificially inseminated honeybee queens. J. Apic. Res., 46(2), 85–95. Google Scholar].
An indicator of the reproductive potential of a queen is, among others, the size of spermatheca and the number of ovarioles in the ovaries. Mean spermathecal volumen in both groups was lower by 0.1 to 0.31 μl when compared to that being found in Carniolan queens by Woyke et al. [1974]Woyke, J., Jasiński, Z., Smagowska, B. (1974). Comparison of reproductive organs and effects of artificial and natural insemination in honey bees of different races and hybrids [Porównanie organów rozrodczych oraz efektów sztucznego unasieniania i naturalnego unasieniania pszczół różnych ras i ich mieszańców]. Pszczeln. Zesz. Nauk., 18, 53–74. Google Scholar, Bieńkowska et al. [2009]Bieńkowska, M., Panasiuk, B., Gerula, D., Węgrzynowicz, P. (2009). Weight of honeybee queens and its effect on the quality of instrumentally inseminated queens. Proceedings of the 4th International Apicultural Congress, Montpellier, France, 15–20 September 2009, p. 135. Google Scholar, Alqarni et al. [2013]Alqarni, A.S., Balhareth, H.M., Owayss, A.A. (2013). Queen morphometric and reproductive characters of Apis mellifera jemenitica, a native honey bee to Saudi Arabia. Bull. Insec., 66(2), 239–244, Google Scholar, and Chuda-Mickiewicz and Samborski [2015]Chuda-Mickiewicz, B., Samborski, J. (2015). The quality of honey bee queens from queen cells incubated at different temperatures. Acta Sci. Pol. Zootechnica, 14(4), 3–10. Google Scholar. The number of ovarioles in the ovary was also lower than that determined in this subspecies by Alqarni et al. [2013]Alqarni, A.S., Balhareth, H.M., Owayss, A.A. (2013). Queen morphometric and reproductive characters of Apis mellifera jemenitica, a native honey bee to Saudi Arabia. Bull. Insec., 66(2), 239–244, Google Scholar, 157, Gregorc and Smodiš Škerl [2015]Gregorc, A., Smodiš Škerl, M.I. (2015). Characteristics of honey bee (Apis mellifera carnica, pollman 1879) queens reared in Slovenian commercial breeding stations. J. Apic. Sci., 59(2), 5–12. https://doi.org/10.1515/jas-2015-0016, 135 to 161, and Chuda-Mickiewicz and Samborski [2015]Chuda-Mickiewicz, B., Samborski, J. (2015). The quality of honey bee queens from queen cells incubated at different temperatures. Acta Sci. Pol. Zootechnica, 14(4), 3–10. Google Scholar, 166 to 178. A similar number of ovarioles in the ovary of Carniolan queens to that determined in our experiment is given by Woyke et al. [1974]Woyke, J., Jasiński, Z., Smagowska, B. (1974). Comparison of reproductive organs and effects of artificial and natural insemination in honey bees of different races and hybrids [Porównanie organów rozrodczych oraz efektów sztucznego unasieniania i naturalnego unasieniania pszczół różnych ras i ich mieszańców]. Pszczeln. Zesz. Nauk., 18, 53–74. Google Scholar, i.e. 125.7.
The lack of significant correlations between the examined physical traits of queens demonstrated in our study coincides with the study by Corbella and Gencalves [1982]Corbella, E., Gencalves, L.S. (1982). Relationship between weight at emergence, number of ovarioles and spermathecal volume of Africanized honeybee queens Apis mellifera. Rev. Brasil. Genet., 5(4), 835–840. Google Scholar, Hatch et al. [1999]Hatch, S., Tarpy, D.R., Fletcher, D.J. (1999). Worker regulation of emergency queen rearing in honey bee colonies and the resultant variation in queen quality. Insectes Sociaux, 46(4), 372–377. https://doi.org/10.1007/s000400050159, and Chuda-Mickiewicz and Samborski [2015]Chuda-Mickiewicz, B., Samborski, J. (2015). The quality of honey bee queens from queen cells incubated at different temperatures. Acta Sci. Pol. Zootechnica, 14(4), 3–10. Google Scholar. They did not find a significant correlation between the body weight of emerging queens and the number of ovarioles and the size of spermatheca. Jockson et at. [2011] demonstrated no correlation between the body weight and the ovariole number, while Chuda-Mickiewicz and Samborski [2015]Chuda-Mickiewicz, B., Samborski, J. (2015). The quality of honey bee queens from queen cells incubated at different temperatures. Acta Sci. Pol. Zootechnica, 14(4), 3–10. Google Scholar between the ovariole number and the spermathecal diameter and volume. The results different from our ones and those of the cited authors were obtained by Kaya et al. [2008], Bieńkowska et al. [2009]Bieńkowska, M., Panasiuk, B., Gerula, D., Węgrzynowicz, P. (2009). Weight of honeybee queens and its effect on the quality of instrumentally inseminated queens. Proceedings of the 4th International Apicultural Congress, Montpellier, France, 15–20 September 2009, p. 135. Google Scholar and, Alqarni et al. [2013]Alqarni, A.S., Balhareth, H.M., Owayss, A.A. (2013). Queen morphometric and reproductive characters of Apis mellifera jemenitica, a native honey bee to Saudi Arabia. Bull. Insec., 66(2), 239–244, Google Scholar who showed a correlation between the body weight and the size of spermatheca. Moreover, Alqarni et al. [2013]Alqarni, A.S., Balhareth, H.M., Owayss, A.A. (2013). Queen morphometric and reproductive characters of Apis mellifera jemenitica, a native honey bee to Saudi Arabia. Bull. Insec., 66(2), 239–244, Google Scholar showed a correlation between the body weight and the number of ovarioles. Woyke [1971]Woyke, J. (1971). Correlation between the age at which honeybee brood was grafted, characteristics of the resultant queens, and result of insemination. J. Apic. Res., 10(1), 45–55. https://doi.org/10.1080/00218839.1971.11099669 also demonstrated significant correlations between physical traits, but only evaluated them in the queens being reared from brood at various ages (eggs, one-day, two-day and three-day larvae) together. The lack of a significant correlation between the body weight of emerging queens and the onset of oviposition being also found in our study is consistent with the study by Szabo et al. [1987]Szabo, T.I., Mills, P.F., Heikel, D.T. (1987). Effects of honeybee queen weight and air temperature on the initiation of oviposition. J. Apic. Res., 26(2), 73–78. https://doi.org/10.1080/00218839.1987.11100740 and Skowronek et al. [2002]Skowronek, W., Kruk, C., Kłopot, J. (2002). Factors affecting oviposition of artificially inseminated honeybee queens. J. Apic. Res., 46(2), 85–95. Google Scholar.
Summing up the obtained results, it can be concluded that the short-term restricted pollen supply to colonies in full season did not affect the quality of reared queen bees and the onset of oviposition.
This study was financed from the statutory activity funds.
Received: 15 Aug 2019
Accepted: 17 Sep 2019
Published online: 3 Dec 2019
Accesses: 1017
Chuda-Mickiewicz, B., Samborski, J., (2019). Effect of restricted pollen supply to colonies on the quality of reared queen bees. Acta Sci. Pol. Zootechnica, 18(3), 21–26. DOI: 10.21005/asp.2019.18.3.04.