, Volume 49, Issue 2, pp 224–234 | Cite as

Upregulation of antioxidant genes in the spermathecae of honey bee (Apis mellifera) queens after mating

  • Alejandra N. Gonzalez
  • Nancy Ing
  • Juliana Rangel
Original article


During storage, the viability of sperm in a honey bee (Apis mellifera) queen’s spermatheca can be decreased by reactive oxygen species. We hypothesized that the expression of antioxidant genes would increase in queen spermathecae after mating. We measured queen morphometric characteristics and expression levels of seven antioxidant-encoding genes in virgin and mated queen spermathecae. We identified a 12% increase in body weight and a fourfold increase in ovary weight in mated queens. There was a twofold higher expression of catalase, thioredoxin 2, and thioredoxin reductase 1 in the spermathecae of mated vs. virgin queens. Expression of the other antioxidant genes (glutathione S-transferase D1, superoxide dismutase 1, vitellogenin, and glyoxalase domain-containing 4-like (GLOD4L) in spermathecae was not different between mated and virgin queens. In drone semen, expression of antioxidant genes was overall low compared to queens except for GLOD4L, which was equivalent to that in queen spermathecae. Increased expression of antioxidant genes may assist in maintaining sperm viability inside the spermathecae of mated queens.


antioxidative enzyme genes Apis mellifera drone honey bee queen spermatheca sperm 



We would like to thank Tammy Olivarez for donating the honey bee queens used in our study. We thank ET Ash and Adrian Fisher II for their assistance in collecting drone samples at the Janice and John G. Thomas Honey Bee Facility at Texas A&M University.

Authors contribution

ANG, NI and JR conceived this research, designed experiments and interpreted the data; ANG performed experiments; ANG, NI and JR performed the analysis, wrote the paper and participated in the revisions of it. All authors read and approved the final manuscript.

Compliance with ethical standards

Conflict of interest

The authors declare that they have no potential conflict of interest in relation to the study in this paper.


  1. Aitken, J., Fisher, H. (1994) Reactive oxygen species generation and human spermatozoa: the balance of benefit and risk. Bioessays 16, 259–67CrossRefPubMedGoogle Scholar
  2. Baer, B. (2005) Sexual selection in Apis bees. Apidologie 36, 187–200CrossRefGoogle Scholar
  3. Baer, B., Eubel, H., Taylor, N.L., O’Toole, N., Millar, A.H. (2009a) Insights into female sperm storage from the spermathecal fluid proteome of the honeybee Apis mellifera. Genome Biol, 10: R67CrossRefPubMedPubMedCentralGoogle Scholar
  4. Baer, B., Heazlewood, J.L., Taylor, N.L., Eubel, H., Millar, A.H. (2009b) The seminal fluid proteome of the honeybee Apis mellifera. Proteomics 9, 2085–2097CrossRefPubMedGoogle Scholar
  5. Berg, S., Koeniger, N., Koeniger, G., Fuchs, S. (1997) Body size and reproductive success of drones (Apis mellifera L). Apidologie 28, 449–460CrossRefGoogle Scholar
  6. Bilodeau, J.F., Blanchette, S., Cormier, N., Sirard, M.A. (2002) Reactive oxygen species-mediated loss of bovine sperm motility in egg yolk Tris extender: protection by pyruvate, metal chelators and bovine liver or oviductal fluid catalase. Theriogenology 57, 1105–22CrossRefPubMedGoogle Scholar
  7. Caron, D.M., Connor, L.J. (2013) Honey Bee Biology and Beekeeping, Revised Edition. Wicwas Press, Kalamazoo, MIGoogle Scholar
  8. Cobey, S.W. (2007) Comparison studies of instrumentally inseminated and naturally mated honey bee queens and factors affecting their performance. Apidologie 38, 390–410.CrossRefGoogle Scholar
  9. Collins, A.M. (2000) Relationship between semen quality and performance of instrumentally inseminated honey bee queens. Apidologie 31, 421–429CrossRefGoogle Scholar
  10. Collins, A.M., Donoghue, A.M. (1999) Viability assessment of honey bee, Apis mellifera, sperm using dual fluorescent staining. Theriogenology 51, 1513–1523CrossRefPubMedGoogle Scholar
  11. Collins, A.M., Williams, V., Evans, J.D. (2004) Sperm storage and antioxidative enzyme expression in the honey bee, Apis mellifera. Insect Mol. Biol. 13, 141–6CrossRefPubMedGoogle Scholar
  12. Collins, A.M., Caperna, T.J., Williams, V., Garrett, W.M., Evans, J.D. (2006) Proteomic analyses of male contributions to honey bee sperm storage and mating. Insect Mol. Biol. 15, 541–549CrossRefPubMedPubMedCentralGoogle Scholar
  13. Corona, M., Robinson, G.E. (2006) Genes of the antioxidant system of the honey bee: annotation and phylogeny. Insect Mol. Biol. 15, 687–701CrossRefPubMedPubMedCentralGoogle Scholar
  14. den Boer, S.P.A., Boomsma, J.J., Baer, B. (2009) Honey bee males and queens use glandular secretions to enhance sperm viability before and after storage. J. Insect Physiol. 55, 538–543CrossRefGoogle Scholar
  15. Du, J., Hincke, M.T., Rose-Martel, M., Hennequet-Antier, C., Brionne, A., Cogburn, L.A., Nys, Y., Gauron, J. (2015) Identifying specific proteins involved in eggshell membrane formation using gene expression analysis and bioinformatics. BMC Genomics 16: 792CrossRefPubMedPubMedCentralGoogle Scholar
  16. Gorshkov, V., Blenau, W., Koeniger, G., Römpp, A., Vilcinskas, A., Spengler, B. (2015) Protein and peptide composition of male accessory glands of Apis mellifera drones investigated by mass spectrometry. PLoS ONE 10:e0125068CrossRefPubMedPubMedCentralGoogle Scholar
  17. Harbo, J.R. (1986) Propagation and instrumental insemination, In: Rinderer T.E. (Ed.), Bee Breeding and Genetics. Academic Press, Inc., Orlando, FL. pp. 361–389CrossRefGoogle Scholar
  18. Heifetz, Y., Rivlin, P.K. (2010) Beyond the mouse model: Using Drosophila as a model for sperm interaction with the female reproductive tract. Theriogenology 73, 723–739CrossRefPubMedGoogle Scholar
  19. Holman, L. (2008) Drosophila melanogaster seminal fluid can protect the sperm of other males. Funct. Ecol. 23, 180–186CrossRefGoogle Scholar
  20. Kanzok, S.M., Fechner, A., Bauer, H., Ulschmid, J.K., Muller, H.M., Botella-Munoz, J.Schneuwly, S., Schrmer, R., Becker, K. (2001) substitution of the thioredoxin system for glutathione reductase in Drosophila melanogaster. Science 291, 643–646CrossRefPubMedGoogle Scholar
  21. King, M., Eubel, H., Millar, A.H., Baer, B. (2011) Proteins within the seminal fluid are crucial to keep sperm viable in the honeybee Apis mellifera. J. Insect Physiol. 57, 409–414CrossRefPubMedGoogle Scholar
  22. Koeniger, G. (1986) Reproduction and mating behavior, in: Rinderer T.E. (Ed.), Bee Breeding and Genetics. Academic Press, Inc., Orlando, FL, pp. 235–252Google Scholar
  23. Koo, H.-N., Lee, S.G., Yun, S.-H., Kim, H.K., Choi, Y.S., Kim G.-H. (2016) Comparative analyses of Cu-Zn superoxide dismutase (SOD1) and thioredoxin reductase (TrxR) at the mRNA level between Apis mellifera L. and Apis cerana F. (Hymenoptera: Apidae) under stress conditions. J. Insect Sci. 16:1–6CrossRefGoogle Scholar
  24. Lapointe, J., Bilodeau, J.F. (2003) Antioxidant defenses are modulated in the cow oviduct during the estrous cycle. Biol. Reprod. 68, 1157–64CrossRefPubMedGoogle Scholar
  25. Malta, J., Martins, G.F., Marques, A.E., Games, P.D., Zanuncio, J.C., Baracat-Pereira, M.C., Salomão, T.M.F. (2014) Insights into the proteome of the spermatheca of the leaf-cutting ant Atta sexdens rubropilosa (Hymenoptera: Formicidae). Fla. Entomol. 97, 1856–1861Google Scholar
  26. Meyer, Y., Buchanan, B.B., Vignols, F., Reichheld, J.-P. (2009) Thioredoxins and glutaredoxins: unifying elements in redox biology. Annu. Rev. Genet. 43, 335–67CrossRefPubMedGoogle Scholar
  27. Monaghan, P., Metcalfe, N.B., Torres, R. (2009) Oxidative stress as a mediator of life history trade-offs: mechanisms, measurements and interpretation. Ecol. Lett. 12, 75–92CrossRefPubMedGoogle Scholar
  28. Munro, D., Treberg, J.R. (2017) A radical shift in perspective: mitochondria as regulators of reactive oxygen species. J Exper. Biol. 220, 1170–1180CrossRefGoogle Scholar
  29. Mustacich, D., Powis, G. (2000) Thioredoxin reductase. Biochem. J. 346, 1–8CrossRefPubMedPubMedCentralGoogle Scholar
  30. Pardini, R.S. (1995) Toxicity of oxygen from naturally occurring redox-active pro-oxidants. Arch. Insect Biochem. 29, 101–118CrossRefGoogle Scholar
  31. Rangel, J., Keller, J. J., Tarpy, D. R. (2012) The effects of honey bee (Apis mellifera L.) queen reproductive potential on colony growth. Insectes Soc. 60, 65–73CrossRefGoogle Scholar
  32. Seehuus, S.C., Norberg, K., Gimsa, U., Krekling, T., Amdam, G.V. (2006) Reproductive protein protects functionally sterile honey bee workers from oxidative stress. Proc. Natl. Acad. Sci. U. S. A. 103, 962–7CrossRefPubMedPubMedCentralGoogle Scholar
  33. Snodgrass, R.E. (1985) Anatomy of the Honey Bee. Cornell University Press, Ithaca, NYGoogle Scholar
  34. Stürup, M., Baer-Imhoof, B., Nash, D.R., Boomsma, J.J., Baer, B. (2013) When every sperm counts: factors affecting male fertility in the honeybee Apis mellifera. Behav. Ecol. 24, 1192–1198CrossRefGoogle Scholar
  35. Svensson, M.J., Larsson, J. (2007) Thioredoxin-2 affects lifespan and oxidative stress in Drosophila. Hereditas 144, 25–32CrossRefPubMedGoogle Scholar
  36. Tarpy, D.R., Nielsen, R., Nielsen, D.I. (2004) A scientific note on the revised estimates of effective paternity frequency in Apis. Insectes Soc. 51, 203–204CrossRefGoogle Scholar
  37. Tavilani, H., Goodarzi, M.T., Vaisi-raygani, A., Salimi, S., Hassanzadeh, T. (2008) Activity of antioxidant enzymes in seminal plasma and their relationship with lipid peroxidation of spermatozoa. Int. Braz. J. Urol. 34, 485–91CrossRefPubMedGoogle Scholar
  38. Verma, L.R., Shuel, R.W. (1973) Respiratory metabolism of the semen of the honey-bee, Apis mellifera. J. Insect Physiol. 19, 97–103CrossRefGoogle Scholar
  39. Weirich, G.F., Collins, A.M., Williams, V.P. (2002) Antioxidant enzymes in the honey bee, Apis mellifera. Apidologie 33, 3–14CrossRefGoogle Scholar
  40. Wilde, J. (1994) Comparison of the development and productivity of bee colonies with naturally and instrumentally inseminated queens kept in different conditions before and after the insemination. Zootechnica 39, 135–152Google Scholar
  41. Winston, M.L. (1987) The Biology of the Honey Bee. Harvard University Press, Cambridge, MAGoogle Scholar

Copyright information

© INRA, DIB and Springer-Verlag France SAS 2017

Authors and Affiliations

  1. 1.Department of EntomologyTexas A&M UniversityCollege StationUSA
  2. 2.Department of Animal ScienceTexas A&M UniversityCollege StationUSA

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