Objectives

• Consider various factors that could influence male mounting behavior
• Design and perform a set of experiments to test whether male mounting behavior is random with respect to sex

Introduction

Bean beetles, Callosobruchus maculatus, spend their short adult life span (1- 2 weeks) mating, with females laying (ovipositing) single fertilized eggs on the surface of various bean species (Talukder & Howse, 1994). Following oviposition, a beetle larva will burrow into the bean and ingest the endosperm portion of the seed before emerging as an adult. The later stages of this process can be readily observed through the appearance of a window where the beetle is pupating. Upon emergence, adults of this species do not require any food or water to survive.

Mating behavior in this species is devoid of courtship and is characterized by a male climbing on a female (mounting) and attempting the transfer of a spermatophore. Mating is also characterized by traumatic insemination of the female due to the male’s spiny genitalia (Crudgington and Siva-Jothy, 2000). Indeed, the female response to male mounting in some cases is to attempt to dismount a male during copulation before a spermatophore is deposited (Eady, 1994), presumably to prevent damage to her reproductive tract (Crudgington and Siva-Jothy, 2000).

Observation of a bean beetle (Callosobruchus maculatus) colony reveals that males, easily identifiable by a lack of white stripe on the caudal end of the abdomen (Figure 1), are regularly seen mounting multiple individuals within the colony, regardless of sex. This interesting animal behavior raises the question of whether or not the mounting is random. More specifically, is male – male mounting an example of intrasexual selection (i.e., competition between males that ultimately leads to greater access to females by one of the males)?

Figure 1. Dorsal view of Callosobruchus maculatus
(From Brown and Downhower, 1988).

Materials

In class, you will be provided with live cultures of bean beetles that have been raised on black eyed peas (Vigna unguiculata). Each culture will contain a dense colony of beetles.  

Experimental Design

You will observe the colonies for occurrences of male mounting behavior for 10 minutes. The groups will then come up with a hypothesis and experimental design to test the significance of this behavior. Then you will perform a collectively decided upon experiment and as a class, we will perform statistical analyses of the data from the experiment.

Each group will then formulate a hypothesis and design an experiment to test possible factors that could influence male mounting behavior. Designs will be presented in a future class meeting and a common experimental design will be selected to test. When designing experiments, you should consider the following:

• List possible advantages and disadvantages for male-male mounting behavior.
• List possible experiences or culture conditions that could influence mounting behavior.
• Describe an experiment to further test whether male-male mounting behavior is random.
• List the variables you would manipulate in your experiment.
• List the variables you would keep constant.
• Predict outcomes for your experiment.

Literature Cited

Beck, C. W. & Blumer, L. S. (2010) A handbook on Bean Beetles, Callosobruchus maculatus. www.beanbeetles.org/handbook/

Chapman T, Arnqvist G, Bangham Brown, L and J.F. Downhower (1988) Analyses in Behavioral Ecology: A Manual of Lab and Field. Sinauer Associates.

Crudgington, H. S. & Siva-Jothy, M. T. (2000) Genital damage, kicking and early death: the battle of the sexes takes a sinister turn in the bean weevil. Nature, 407, 855–856.

Eady, P.E. (1994) Intraspecific variation in sperm precedence in the bruchid beetle Callosobruchus maculatus. Ecological Entomology 19: 11-16.

Maklakov, A. A. & R. Bonduriansky (2009) Sex differences in survival costs of homosexual and heterosexual interactions: evidence from a fly and a beetle. Anim. Behav. 77:1375–1379.

Talukder FA & Howse PE. (1994) Repellent, toxic and food protectant effects of pithraj, Aphanamixis polystachya extracts against the pulse beetle, Callosobruchus chinensis in storage. Journal of Chemical Ecology 20:899-908.

Thornhill, R. & Alcock, J. (1983) The Evolution of Insect Mating Systems. Cambridge, Massachusetts: Harvard University Press.

This experiment was written by M. Clark and L. Blumke.  

Copyright © by Merry Clark and Lisa Blumke, 2012. All rights reserved. The content of this site may be freely used for non-profit educational purposes, with proper acknowledgement of the source. All other uses are prohibited without prior written permission from the copyright holders.

In this activity, students observe a specific social behavior in male Callosobruchus maculatus. This lab exercise allows students to consider the reasons for this behavior as well as how environmental, developmental, or social environments could affect it. Student groups perform an introductory experiment to test the frequency of mounting attempts made by males, either toward females or other males. The class then performs statistical tests to determine if there is a significant difference between male-male versus male-female mounting behavior. As a class, we discuss possible factors that could influence male mounting, and students are asked to (go home and) design an experiment to test these possibilities. In a follow-up class, students present their experimental designs and (as a class) we select one to perform. Since the follow up experiment occurs during a subsequent class meeting, preexisting colonies of various types will be required (see "Equipment and supplies").

Experimental designs may involve tracking male mounts by marking one or more males in a colony. We found that males can be easily marked with a spot of white paint on the dorsal surface without the need to chill them.

Experimental Design

In the first laboratory class, students observe dense cultures of bean beetles and observe any bean beetle demonstrating mounting behavior, regardless of sex. Students then design an experiment to test the statistical significance of this behavior and determine whether male mounting behavior is random. The class then discusses what factors might influence the randomness, or lack of randomness, of male-male mounting. After student group and class discussions, students independently formulate hypotheses and design experiments to explain male mounting behavior. In a second class, after students briefly present their designs, a single experiment is chosen to further investigate male mounting behavior.

Questions you might expect students to ask:

• Does the ratio of males to females influence male mounting behavior?
• Does the reproductive experience of the male influence male mounting behavior?
• Does the reproductive experience of the female being mounted influence male mounting behavior?
• Does the reproductive experience of the male being mounted influence male mounting behavior?
• Is a mounted male affected, behaviorally or physiologically, by being mounted?
• Do culture conditions (bean type, bean density, temperature, etc.) influence male mounting behavior?

Data Collection

For a typical experiment, students will record the number of times a male mounts a female versus a male. The data will be pooled and analyzed using binomial tests to compare frequency of mounts. Additional analysis may be needed depending on the follow up experiment.

A single class, usually 6 -8 groups of 3 or 4 students, should provide enough replicates to be sufficient. Data can be collected in simple table form, with tallies or checks, to record each observed mounting. Time frames could also be added to track the amount of time spent mounting, spent in the mounting position, or incrementally over a given time frame. Data collection can be collected the same day the experiment is done or later, as dictated by the experiment.

Data Analysis

For a typical experiment, students will record the number of times a male mounts a female versus a male. The data will be pooled and analyzed using binomial tests to compare frequency of mounts. Additional analysis may be needed depending on the follow up experiment.

Equipment and Supplies

For a class of 24 students working in groups of 4:

• 6 bean beetle cultures (150mm plates with monolayer of black eyed peas), with dense colonies of beetles
• White, non-toxic acrylic paint
• 6 handheld magnifying glasses

 

This experiment was written by M. Clark and L. Blumke.

Copyright © by Merry Clark and Lisa Blumke , 2012. All rights reserved. The content of this site may be freely used for non-profit educational purposes, with proper acknowledgement of the source. All other uses are prohibited without prior written permission from the copyright holders.

Students observed plates of bean beetle colonies, containing ratios of 5:5, 10:10, or 15:15 male to female beetles. A marked male was tracked by multiple students within a single group. The number of times that the marked male was seen mounting any other beetle was recorded in a tally format. After 10 minutes, the same plates were redistributed to different groups and another tracking period began. This method was continued for the duration of 1 hour. After all the data were collected (Table 1), we performed binomial tests to compare the frequency of male-male versus male-female mounts.

The ratio of male to female beetles within a colony had a marked effect on the randomness of mounting behavior observed when the data were pooled across trials, but not within individual trials (Table 1). Colonies of 5:5 males:females exhibited random mounting behavior. In contrast, colonies with larger number of beetles showed a bias toward male-female mounting behavior.

Table 1: The effect of density on male mounting behavior

Student Handout [pdf] [docx]

Instructor's Notes [pdf] [docx]

Sample data table [pptx]

Sample data [xlsx]

Identifying the sexes [ppt]