Behavioral Ecology and Sociobiology

, Volume 70, Issue 4, pp 519–532 | Cite as

Multiple mechanisms of cryptic female choice act on intraspecific male variation in Drosophila simulans

  • Outi Ala-Honkola
  • Mollie K. Manier
Original Article


Postcopulatory sexual selection can arise when females mate with multiple males and is usually mediated by an interaction between the sexes. Cryptic female choice (CFC) is one form of postcopulatory sexual selection that occurs when female morphology, physiology, or behavior generates a bias in fertilization success. However, its importance in nonrandom reproductive success is poorly resolved due to challenges distinguishing the roles of females and males in generating patterns of fertilization bias. Nevertheless, two CFC mechanisms have recently been documented and characterized in Drosophila simulans within the context of gametic isolation in competitive hybrid matings with Drosophila mauritiana: sperm ejection and nonrandom use of sperm storage organs for fertilization. Here, we explore if and how female D. simulans employ these two mechanisms of CFC in response to intraspecific male size variation. We used transgenic males expressing green (GFP) or red fluorescent protein (RFP) in sperm heads to document postcopulatory processes, in conjunction with a probabilistic analytical model. We unexpectedly found that differential reproductive success was also a function of male population (GFP or RFP), suggesting that females use different CFC mechanisms to select for different male traits. Moreover, concordance of selection at the precopulatory (as measured by mating latency) and postcopulatory stages depends on both the male trait and the CFC mechanism examined. Larger males were more successful both before and after mating, but we unexpectedly found that females also mated more quickly with males with GFP-labeled sperm, while fertilization bias favored RFP-labeled sperm.


Precopulatory sexual selection Postcopulatory sexual selection Sperm competition Female ejection Female preference Fertilization bias 



Scott Pitnick provided laboratory space, equipment, and valuable discussions, while Liz O’Hanlon assisted with experiments.

Compliance with ethical standards


The work was supported by two National Science Foundation grants to Manier (DEB-1145965 and DEB-1257859) and an Academy of Finland grant to Ala-Honkola (grant 250999).

Conflict of interest

Authors Ala-Honkola and Manier declare they have no conflicts of interest.

Ethical approval

All applicable international, national, and/or institutional guidelines for the care and use of animals were followed. This article does not contain any studies with human participants performed by any of the authors.

Supplementary material

265_2016_2069_MOESM1_ESM.docx (268 kb)
ESM 1 (DOCX 268 kb)


  1. Akaike H (1973) Information theory as an extension of the maximum likelihood principle. In: Petrov BN, Csaki F (eds) Second international symposium on information theory. Akadémiai Kiadó, Budapest, pp 267–281Google Scholar
  2. Ala-Honkola O, Manier MK, Lüpold S, Pitnick S (2011) No evidence for postcopulatory inbreeding avoidance in Drosophila melanogaster. Evolution 65:2699–2705. doi: 10.1111/j.1558-5646.2011.01317.x CrossRefPubMedGoogle Scholar
  3. Amitin EG, Pitnick S (2007) Influence of developmental environment on male- and female-mediated sperm precedence in Drosophila melanogaster. J Evol Biol 20:381–391. doi: 10.1111/j.1420-9101.2006.01184.x CrossRefPubMedGoogle Scholar
  4. Anderson MJ, Dixson AS, Dixson AF (2006) Mammalian sperm and oviducts are sexually selected: evidence for co-evolution. J Zool 270:682–686. doi: 10.1111/j.1469-7998.2006.00173.x CrossRefGoogle Scholar
  5. Bangham J, Chapman T, Partridge L (2002) Effects of body size, accessory gland and testis size on pre- and postcopulatory reproductive success in Drosophila melanogaster. Anim Behav 64:915–921. doi: 10.1006/anbe.2002.1976 CrossRefGoogle Scholar
  6. Birkhead TR, Møller AP, Sutherland WJ (1993) Why do females make it so difficult for males to fertilize their eggs? J Theor Biol 161:51–60. doi: 10.1006/jtbi.1993.1039 CrossRefGoogle Scholar
  7. Brennan PLR, Prum RO, McCracken KG, Sorenson MD, Wilson RE, Birkhead TR (2007) Coevolution of male and female genital morphology in waterfowl. PLoS One 5, e418. doi: 10.1371/journal.pone.0000418 CrossRefGoogle Scholar
  8. Bretman A, Newcombe D, Tregenza T (2009) Promiscuous females avoid inbreeding by controlling sperm storage. Mol Ecol 18:3340–3345. doi: 10.1111/j.1365-294X.2009.04301.x CrossRefPubMedGoogle Scholar
  9. Byrne PG, Rice WR (2005) Remating in Drosophila melanogaster: an examination of the trading-up and intrinsic male-quality hypotheses. J Evol Biol 18:1324–1331. doi: 10.1111/j.1420-9101.2005.00918.x CrossRefPubMedGoogle Scholar
  10. Candolin U (2003) The use of multiple cues in mate choice. Biol Rev 78:575–595. doi: 10.1017/S1464793103006158 CrossRefPubMedGoogle Scholar
  11. Clark AG, Begun DJ (1998) Female genotypes affect sperm displacement in Drosophila. Genetics 149:1487–1493PubMedPubMedCentralGoogle Scholar
  12. Clark AG, Begun DJ, Prout T (1999) Male × female interactions in Drosophila sperm competition. Science 283:217–220. doi: 10.1126/science/283.5399.217 CrossRefPubMedGoogle Scholar
  13. Crawley MJ (2007) The R book. Wiley, West SussexCrossRefGoogle Scholar
  14. Danielsson I (2001) Antagonistic pre- and postcopulatory sexual selection on male body size in a water strider (Gerris lacustris). Proc R Soc Lond B 268:77–81. doi: 10.1098/rspb.2000.1332 CrossRefGoogle Scholar
  15. Dean R, Nakagawa S, Pizzari T (2011) The risk and intensity of sperm ejection in female birds. Am Nat 178:343–354. doi: 10.1086/661244 CrossRefPubMedGoogle Scholar
  16. Droge-Young EM, Manier MK, Lüpold S, Belote JM, Pitnick S (2012) Covariance among premating, post-copulatory, and viability fitness components of Drosophila melanogaster and their influence on paternity measurement. J Evol Biol 25:1555–1563. doi: 10.1111/j.1420-9101.2012.02540.x CrossRefPubMedGoogle Scholar
  17. Dybas LK, Dybas HS (1981) Coadaptation and taxonomic differentiation of sperm and spermathecae in featherwing beetles. Evolution 35:168–174CrossRefGoogle Scholar
  18. Eberhard WG (1985) Sexual selection and animal genitalia. Harvard University Press, Cambridge. ISBN 0-674-80283-7CrossRefGoogle Scholar
  19. Eberhard WG (1996) Female control: sexual selection by cryptic female choice. Princeton University, Princeton. ISBN 0-691-01084-6Google Scholar
  20. Edvardsson M, Arnqvist G (2000) Copulatory courtship and cryptic female choice in red flour beetles Tribolium castaneum. Proc R Soc Lond B 267:559–563. doi: 10.1098/rspb.2000.1037 CrossRefGoogle Scholar
  21. Engqvist L, Dekomien G, Lippmann T, Epplen JT, Sauer KP (2007) Sperm transfer and paternity in the scorpionfly Panorpa cognata: large variance in traits favoured by postcopulatory episodes of sexual selection. Ecol Evol 21:801–816. doi: 10.1007/s10682-006-9152-6 CrossRefGoogle Scholar
  22. Evans JP, Marshall DJ (2005) Male-by-female interactions influence fertilization success and mediate the benefits of polyandry in the sea urchin Heliocidaris erythrogramma. Evolution 59:106–112CrossRefPubMedGoogle Scholar
  23. Evans JP, Zane L, Francescato S, Pilastro A (2003) Directional postcopulatory sexual selection revealed by artificial insemination. Nature 421:360–363. doi: 10.1038/nature01367 CrossRefPubMedGoogle Scholar
  24. Evans JP, Rosengrave P, Gasparini C, Gemmell N (2013) Delineating the roles of males and females in sperm competition. Proc R Soc Lond B 280:20132047. doi: 10.1098/rspb.2013.2047 CrossRefGoogle Scholar
  25. Fricke C, Martin OY, Bretman A, Bussiere LF, Chapman T (2010) Sperm competitive ability and indices of lifetime reproductive success. Evolution 64:2746–2757. doi: 10.1111/j.1558-5646.2010.01022.x CrossRefPubMedGoogle Scholar
  26. Hellriegel B, Bernasconi G (2000) Female-mediated differential sperm storage in a fly with complex spermathecae, Scatophaga stercoraria. Anim Behav 59:311–317. doi: 10.1006/anbe.1999.1308 CrossRefPubMedGoogle Scholar
  27. Hellriegel B, Ward PI (1998) Complex female reproductive tract morphology: its possible use in postcopulatory female choice. J Theor Biol 190:179–186. doi: 10.1006/jtbi.1997.0546 CrossRefGoogle Scholar
  28. Higginson DM, Miller KB, Segraves KA, Pitnick S (2012) Female reproductive tract form drives the evolution of complex sperm morphology. PNAS 109:4538–4543. doi: 10.1073/pnas.1111474109 CrossRefPubMedPubMedCentralGoogle Scholar
  29. Hosken DJ, Taylor ML, Hoyle K, Higgins S, Wedell N (2008a) Attractive males have greater success in sperm competition. Curr Biol 18:R553–R554. doi: 10.1016/j.cub.2008.04.028 CrossRefPubMedGoogle Scholar
  30. Hosken DJ, Taylor ML, Hoyle K, Higgins S, Wedell N (2008b) Attractive males have greater success in sperm competition. Curr Biol 18:R553–R554CrossRefPubMedGoogle Scholar
  31. Hothorn T, Bretz F, Westfall P (2008a) Simultaneous inference in general parametric models. Biom J 50:346–363. doi: 10.1002/bimj.200810425 CrossRefPubMedGoogle Scholar
  32. Hothorn T, Bretz F, Westfall P, Heiberger RM (2008) multcomp: simultaneous Inference in general parametric models. URL:
  33. House CM, Lewis Z, Hodgson DJ, Wedell N, Sharma MD, Hunt J, Hosken DJ (2013) Sexual and natural selection both influence male genital evolution. PLoS One 8, e63807. doi: 10.1371/journal.pone.0063807 CrossRefPubMedPubMedCentralGoogle Scholar
  34. Ingleby FC, Hunt J, Hosken DJ (2013) Genotype-by-environment interactions for female mate choice of male cuticular hydrocarbons in Drosophila simulans. PLoS One 8, e67623. doi: 10.1371/journal.pone.0067623 CrossRefPubMedPubMedCentralGoogle Scholar
  35. Katayama N, Abbott JK, Kjærandsen J, Takahashi Y, Svensson EI (2014) Sexual selection on wing interference patterns in Drosophila melanogaster. PNAS 111:15144–15148. doi: 10.1073/pnas.1407595111 CrossRefPubMedPubMedCentralGoogle Scholar
  36. Lehtonen TK, Rintakoski S, Lindström K (2007) Mate preference for multiple cues: interplay between male and nest size in the sand goby, Pomatoschistus minutus. Behav Ecol 18:696–700. doi: 10.1093/beheco/arm032 CrossRefGoogle Scholar
  37. Lewis SM, Austad SN (1990) Sources of intraspecific variation in sperm precedence in red flour beetles. Am Nat 135:351–359. doi: 10.1086/285050 CrossRefGoogle Scholar
  38. Lewis SM, Austad SN (1994) Sexual selection in flour beetles: the relationship between sperm precedence and male olfactory attractiveness. Behav Ecol 5:219–224CrossRefGoogle Scholar
  39. Locatello L, Rasotto MB, Evans JP, Pilastro A (2006) Colourful male guppies produce faster and more viable sperm. J Evol Biol 19:1595–1602. doi: 10.1111/j.1420-9101.2006.01117.x CrossRefPubMedGoogle Scholar
  40. Lüpold S, Manier MK, Ala-Honkola O, Belote JM, Pitnick S (2011) Male Drosophila melanogaster adjust ejaculate size based on female mating status, fecundity, and age. Behav Ecol 22:184–191. doi: 10.1093/beheco/arq193 CrossRefGoogle Scholar
  41. Lüpold S, Manier MK, Berben KS, Smith KJ, Daley BD, Buckley SH, Belote JM, Pitnick S (2012) How multivariate ejaculate traits determine competitive fertilization success in Drosophila melanogaster. Curr Biol 22:1667–1672. doi: 10.1016/j.cub.2012.06.059 CrossRefPubMedGoogle Scholar
  42. Lüpold S, Pitnick S, Berben KS, Blegnini CS, Belote JM, Manier MK (2013) Female mediation of competitive fertilization success in Drosophila melanogaster. PNAS 110:10693–10698. doi: 10.1073/pnas.1300954110 CrossRefPubMedPubMedCentralGoogle Scholar
  43. Manier MK, Belote JM, Berben KS, Novikov D, Stuart WT, Pitnick S (2010) Resolving mechanisms of competitive fertilization success in Drosophila melanogaster. Science 328:354–357. doi: 10.1126/science.1187096 CrossRefPubMedGoogle Scholar
  44. Manier MK, Belote JM, Berben KS, Lüpold S, Ala-Honkola O, Collins WF, Pitnick S (2013a) Rapid diversification of sperm precedence traits and processes among three sibling Drosophila species. Evolution 67:2348–2362. doi: 10.1111/evo.12117 CrossRefPubMedGoogle Scholar
  45. Manier MK, Lüpold S, Belote JM, Starmer WT, Berben KS, Ala-Honkola O, Collins WF, Pitnick S (2013b) Postcopulatory sexual selection generates speciation phenotypes in Drosophila. Curr Biol 23:1853–1862. doi: 10.1016/j.cub.2013.07.086 CrossRefPubMedGoogle Scholar
  46. Manier MK, Lüpold S, Pitnick S, Starmer WT (2013c) An analytical framework for estimating fertilization bias and the fertilization set from multiple sperm-storage organs. Am Nat 182:552–561. doi: 10.1086/671782 CrossRefPubMedGoogle Scholar
  47. Markow TA, Ricker JP (1992) Male size, developmental stability, and mating success in satural populations of three Drosophila species. Heredity 69:122–127. doi: 10.1038/hdy.1992.104 CrossRefPubMedGoogle Scholar
  48. Miller GA, Pitnick S (2002) Sperm-female coevolution in Drosophila. Science 298:1230–1233. doi: 10.1126/science.1076968 CrossRefPubMedGoogle Scholar
  49. Minder AM, Hosken DJ, Ward PI (2005) Co-evolution of male and female reproductive characters across the Scathophagidae (Diptera). J Evol Biol 18:60–69. doi: 10.1111/j.1420-9101.2004.00799.x CrossRefPubMedGoogle Scholar
  50. Morrow EH, Arnqvist G (2003) Costly traumatic insemination and a female counter-adaptation in bed bugs. Proc R Soc Lond B 270:2377–2381. doi: 10.1098/rspb.2003.2514 CrossRefGoogle Scholar
  51. Nilsson T, Fricke C, Arnqvist G (2003) The effects of male and female genotype on variance in male fertilization success in the red flour beetle (Tribolium castaneum). Behav Ecol Sociobiol 53:227–233. doi: 10.1007/s00265-002-0565-0 CrossRefGoogle Scholar
  52. Parker GA (1970) Sperm competition and its evolutionary consequences in the insects. Biol Rev 45:525–567. doi: 10.1111/j.1469-185X.1970.tb01176.x CrossRefGoogle Scholar
  53. Parker GA, Simmons LW, Kirk H (1990) Analysing sperm competition data: simple models for predicting mechanisms. Behav Ecol Sociobiol 27:55–65CrossRefGoogle Scholar
  54. Partridge L, Holliday T (1984) Mating patterns and mate choice. In: Krebs JR, Davies NB (eds) Behavioural ecology: an evolutionary approach, 2nd edn. Sinauer Associates, Sunderland, pp 222–250. ISBN 0-87893-133-3Google Scholar
  55. Partridge L, Ewing A, Chandler A (1987) Male size and mating success in Drosophila melanogaster: the roles of male and female behavior. Anim Behav 35:555–562CrossRefGoogle Scholar
  56. Pilastro A, Simonato M, Bisazza A, Evans JP (2004) Cryptic female preference for colorful males in guppies. Evolution 58:665–669CrossRefPubMedGoogle Scholar
  57. Pischedda A, Rice WR (2012) Partitioning sexual selection into its mating success and fertilization success components. PNAS 109:2049–2053. doi: 10.1073/pnas.1110841109 CrossRefPubMedPubMedCentralGoogle Scholar
  58. Pitnick S (1991) Male size influences mate fecundity and remating interval in Drosophila melanogaster. Anim Behav 41:735–745. doi: 10.1016/S0003-3472(05)80340-9 CrossRefGoogle Scholar
  59. Pitnick S, Brown WD (2000) Criteria for demonstrating female sperm choice. Evolution 54:1052–1056CrossRefPubMedGoogle Scholar
  60. Pitnick S, Garcia-González F (2002) Harm to females increases with male body size in Drosophila melanogaster. Proc R Soc Lond B 269:1821–1828. doi: 10.1098/rspb.2002.2090 CrossRefGoogle Scholar
  61. Pitnick S, Markow TA, Spicer GS (1999) Evolution of multiple kinds of female sperm-storage organs in Drosophila. Evolution 53:1804–1822CrossRefGoogle Scholar
  62. Pitnick S, Miller GA, Schneider K, Markow TA (2003) Ejaculate-female coevolution in Drosophila mojavensis. Proc R Soc Lond B 270:1507–1512. doi: 10.1098/rspb.2003.2382 CrossRefGoogle Scholar
  63. Pizzari T, Birkhead TR (2000) Female feral fowl eject sperm of subdominant males. Nature 405:787–789. doi: 10.1038/35015558 CrossRefPubMedGoogle Scholar
  64. Polak M, Simmons LW (2009) Secondary sexual trait size reveals competitive fertilization success in Drosophila bipectinata Duda. Behav Ecol 20:753–760. doi: 10.1093/beheco/arp056 CrossRefGoogle Scholar
  65. Presgraves DC, Baker RH, Wilkinson GS (1999) Coevolution of sperm and female reproductive tract morphology in stalk-eyed flies. Proc R Soc Lond B 266:1041–1047CrossRefGoogle Scholar
  66. R Development Core Team (2013) R: a language and environment for statistical computing. R Foundation for Statistical Computing, ViennaGoogle Scholar
  67. Ritchie MG, Halsey EJ, Gleason JM (1999) Drosophila song as a species-specific mating signal and the behavioural importance of Kyriacou and Hall cycles in D. melanogaster song. Anim Behav 58:649–657. doi: 10.1006/anbe.1999.1167 CrossRefPubMedGoogle Scholar
  68. Robertson FW, Reeve E (1952) Studies in quantitative inheritance. I. The effects of selection on wing and thorax length in Drosophila melanogaster. J Genet 50:414–448. doi: 10.1007/BF02986839 CrossRefGoogle Scholar
  69. Rönn JL, Katvala M, Arnqvist G (2011) Correlated evolution between male and female primary reproductive characters in seed beetles. Funct Ecol 25:634–640. doi: 10.1111/j.1365-2435.2010.01809.x CrossRefGoogle Scholar
  70. Rosengrave P, Gemmell NJ, Metcalf V, McBride K, Montgomerie R (2008) A mechanism for cryptic female choice in chinook salmon. Behav Ecol 19:1179–1185. doi: 10.1093/beheco/arn089 CrossRefGoogle Scholar
  71. Rugman-Jones PF, Eady PE (2007) Conspecific sperm precedence in Callosobruchus subinnotatus (Coleoptera: Bruchidae): mechanisms and consequences. Proc R Soc Lond B 274:983–988. doi: 10.1098/rspb.2006.0343 CrossRefGoogle Scholar
  72. Rugman-Jones PF, Eady PE (2008) Co-evolution of male and female reproductive traits across the Bruchidae (Coleoptera). Funct Ecol 22:880–886. doi: 10.1111/j.1365-2435.2008.01446.x CrossRefGoogle Scholar
  73. Sakaluk SK, Eggert A-K (1996) Female control of sperm transfer and intraspecific variation in sperm precedence: antecedents to the evolution of a courtship food gift. Evolution 50:694–703CrossRefGoogle Scholar
  74. SAS Institute (2008) SAS/STAT 9.2 user’s guide, 2nd edn. SAS Institute, CaryGoogle Scholar
  75. Sharma MD, Hunt J, Hosken DJ (2012) Antagonistic responses to natural and sexual selection and the sex-specific evolution of cuticular hydrocarbons in Drosophila simulans. Evolution 66:665–677. doi: 10.1111/j.1558-5646.2011.01468.x CrossRefPubMedGoogle Scholar
  76. Simmons LW, Parker GA, Stockley P (1999) Sperm displacement in the yellow dung fly, Scatophaga stercoraria: an investigation of male and female processes. Am Nat 153:302–314. doi: 10.1086/303171 CrossRefGoogle Scholar
  77. Simmons LW, Thomas ML, Simmons FW, Zuk M (2013) Female preferences for acoustic and olfactory signals during courtship: male crickets send multiple messages. Behav Ecol 24:1099–1107. doi: 10.1093/beheco/art036 CrossRefGoogle Scholar
  78. Siva-Jothy MT (1987) Variation in copulation duration and the resultant degree of sperm removal in Orthetrum cancellatum (L.) (Libellulidae: Odonata). Behav Ecol Sociobiol 20:147–151. doi: 10.1007/BF00572637 CrossRefGoogle Scholar
  79. Snook RR, Hosken DJ (2004) Sperm death and dumping in Drosophila. Nat 428:939–941Google Scholar
  80. Taylor ML, Wedell N, Hosken DJ (2008) Sexual selection and female fitness in Drosophila simulans. Behav Ecol Sociobiol 62:721–728. doi: 10.1007/s00265-007-0497-9 CrossRefGoogle Scholar
  81. Thornhill R (1983) Cryptic female choice and its implications in the scorpionfly Harpobittacus nigriceps. Am Nat 122:765–788. doi: 10.1086/284170 CrossRefGoogle Scholar
  82. Urbach D, Folstad I, Rudolfsen G (2005) Effects of ovarian fluid on sperm velocity in Arctic charr (Salvelinus alpinus). Behav Ecol Sociobiol 57:438–444. doi: 10.1007/s00265-004-0876-4 CrossRefGoogle Scholar
  83. Venables WN, Ripley BD (2002) Modern applied statistics with S, 4th edn. Springer, New York. ISBN 978-0-387-21706-2CrossRefGoogle Scholar
  84. Vortman Y, Lotem A, Dor R, Lovette I, Safran RJ (2013) Multiple sexual signals and behavioral reproductive isolation in a diverging population. Am Nat 182:514–523. doi: 10.1086/671908 CrossRefPubMedGoogle Scholar
  85. Zuur AF, Hilbe JM, Ieno EN (2013) A beginner’s Guide to GLM and GLMM with R: a frequentist and Bayesian perspective to ecologists. Highland Statistics Ltd., Newburgh. ISBN 978-0-387-93837-0Google Scholar

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© Springer-Verlag Berlin Heidelberg 2016

Authors and Affiliations

  1. 1.Department of Biological and Environmental ScienceUniversity of JyvaskylaJyvaskylaFinland
  2. 2.Biological SciencesThe George Washington UniversityWashingtonUSA

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