Research Project

Effects of Female Appearance on Male Mating Behavior in Danaus plexippus

Erin Layde, Kate Aughenbaugh, Ulysses Izaola, Cristina Flores, Michelle O’Connell, Louis Baheriy
Humboldt Middle School
St. Paul, MN
April 2000

Abstract

We investigated how the characteristics of a female monarch butterfly affected mate recognition by male monarchs. The specific characteristic that we studied was appearance, namely color and pattern. We discovered that male monarchs use color, specifically orange, to recognize another monarch as a potential mate. Other factors, such as gender and appearing like a male, did not discourage male monarchs from mating attempts. Our results confirm that males do use appearance to recognize females as potential mates. Our results indicate that orange coloration is the most significant characteristic that males use to find mating partners.

Introduction

We explored how the appearance of a female monarch butterfly affects a male’s ability to recognize it as a potential mate. We became interested in this question because prior research and observations have shown that male monarchs will attempt to mate with other males, dead monarchs, and falling leaves. In a mating study conducted in the overwintering sites in Mexico, 25-28% of mating attempts were between two males, and male-male attempts were as long as male-female attempts (Oberhauser, 1999). Although most relatives of the monarch butterfly use pheromones to locate potential mates, chemical cues seem to be unnecessary for monarchs (Alcock, 1998). It is a bit suprising, however, that monarchs seem to be so indiscriminant about their mating choice because, in general, male species seem to be choosy about their mates when mating involves passing on valuable goods to females (Alcock, 1998). Because male monarchs pass on their valuable spermatophore to females during mating, it is likely that male monarchs use some cue to determine the best mate. In this experiment, we tested to see if physical appearance was that cue. We became interested in how the appearance affects mating choice because it is known that monarchs are highly dependent upon vision.

Hypotheses

Ho: Appearance has no effect on the males’ ability to recognize females as potential mates.
Ha: Appearance has an effect on males’ ability to recognize females as potential mates.

Methods

Our experimental design consisted of observing the mating behavior of male monarch butterflies while in a cage with female monarch butterflies whose appearance had been changed. We obtained some female monarchs with a natural color variation and manipulated others by changing their color and making some appear male.

We obtained 16 female and 8 male monarch butterflies from the University of Minnesota, Cornell Stock, an inbred line. We used 4 different female appearance categories: "white", "black dotted", and "regular". Four of the female monarch butterflies had a mutation causing them to have white or pale orange scales instead of the normal bright orange coloration. Using a black permanent marker, we altered the appearance of four female monarchs by coloring their wings all black. We altered four more females by adding a black dot to their hind wings to mimic male monarchs. We left the males and the last four females unaltered. We then placed all of the monarchs, male and female, into an outdoor 4’x 4’x 8’ screen cage. We observed and cataloged behavior for two hours on a sunny day. Based on these observations, we identified the following behaviors as male mating behaviors:

Chase: A male monarch chases a female monarch for over 1 meter for 5 seconds or more.
Land-on: A male monarch lands directly on top of another monarch and remains there for at least 5 seconds.
Attempts: A male monarch stations itself on a monarch for at least 30 seconds and can be visibly seen trying to connect abdomens.
Mate: A male monarch successfully mates with a female monarch and remains in that position until morning.

We observed and recorded the males’ mating behaviors on 6 consecutive days in July 1999, for 2 hours each day. We recorded the behaviors, chase, land- on, attempt, and mate, each time the behavior occurred. We used a chi-square test to test for an association between male mating behavior frequency and butterfly appearance.

Results

Male monarchs ignored the "black" females, directing zero mating behaviors toward them. Males also directed few mating behaviors toward "white" females. Regular females experienced more mating behaviors from the males than any other group. Males directed approximately the same number of mating behaviors toward "black dotted" female monarchs and other male monarchs (Figure 1).

We also categorized the butterflies by color: white, black, and orange. Orange butterflies include "regular" females, "black dotted" females, and males. Male mating behavior was significantly associated with butterfly color; males directed a higher than expected number of mating behaviors toward orange butterflies (chi-square = 21.47, p<0.005). These results took into account that there were sixteen orange butterflies compared to only four white and four black butterflies (Table 1 and Figure 2). We can, therefore, reject our null hypothesis that appearance has no effect on male monarchs’ ability to recognize potential mates.

Figure 1: No mating behaviors were performed on black females, 9 behaviors were performed on white females, 33 mating behaviors were performed on the dotted females, 45 mating behaviors on the "normal" females and 31 mating behaviors were performed on every 4 males.
* Number was divided by 2 because there were twice as many males as other female monarch types.

Figure 2: No mating behavior was performed on the black butterflies, 9 mating behaviors were performed on the white butterflies while 70.25* mating behaviors were performed on average of 4 orange butterflies.
* Graph takes into account the fact that there were 4 times as many orange monarchs than other types.

Chi-square test: We used chi-square tests to determine the significance of appearance on male mating behavior. To calculate the expected values, we multiplied the total number of observed attempts by the proportion of butterflies in each category.

Table 1a.
Total of Each Color	16 orange	4 white	4 black
Percentage	66%	17%	17%

Table 1b.
	white	black	orange	Total
Observed Attempts	2	0	41	43
Expected Attempts	7.31	7.31	28.38	43

Statistical chi-squares for orange white and black colors:
Chase: x cal 24.17> x crit 10.59 therefore reject Ho @99.5 confidence level.

Discussion

Our results show that appearance, specifically color, does significantly affect the mating behavior of males. More detailed aspects of appearance, such as presence of black dots, seem to be less important in mate recognition. Males seemed to be just as interested in mating with males as they were with orange females (Figure 2).

During our experiment, black monarchs were less active during the last three days of our testing period compared to the other monarchs. Their wings had greater damage than the other monarchs and there was a greater potential for leftover permanent marker scent because more marker was used on their wings compared to other monarchs used in our experiment. It is possible that their potential permanent marker scent and their low activity level could have caused some of the lack of interest in the black monarchs during the last three days of the experiment.

If we were to further explore our question, we would color the monarchs using a method that did not alter their scent or wing condition. It would also be interesting to investigate how males respond to other species of orange butterflies such as the Viceroy. Other areas of interest include how the movement and scent of females affect males’ mating behavior.

Acknowledgments

We would like to thank Karen Oberhauser and Michelle Prysby for their guidance and support throughout the research project. The National Science Foundation and The Monarchs in the Classroom Program at the University of Minnesota supported our research. Both programs provided us with our experimental specimens, monarch adults. We would also like to thank all of the dedicated staff from the Monarch Monitoring Project who taught us the research skills necessary to make our experiment a success. A final thanks to all of the parents who provided transportation and supported our project.

Literature Cited

Alcock, John. 1998. Animal Behavior: an evolutionary approach. Pg. 429.

Oberhauser, K.S. and D. Frey. 1999. Coerced Mating in Monarch Butterflies. Proceedings of the North American Conference on the Monarch Butterfly, p. 67-68. I. Pisanty, K. Oberhauser, L. Merino, and S. Price, editors.