Common Behaviours observed in the Seed Beetle Callosobruchus maculatus

An observational study to determine the common behaviours of Seed beetles carried out across five foundation study groups

Objective

Using the seed beetle Callosobruchus maculatus, we aimed to understand what common behaviours male and female beetles exhibited when introduced as pairs within a glass container for approximately 45 minutes. C. maculatus is a common pest of stored legumes and are easy to rear for studies to understand their life-cycle. Larvae (four larval instars) of this species burrow into the dry seeds and remain there by feeding on the seed for about 20 days (temperature dependent). Once they pupate they emerge from the seed as a mature adult and can readily mate with females. Females of this species (and strain) are known to be bigger than males and each female can lay up to 200 eggs in their lifetime (condition dependent). Males and females can be distinguished by their patterns in their elytron- females have a distinct box like appearance on their dorsal surface and males generally lack them and have a plain elytron, however, some males seem to have these markings too. A more effective way of distinguishing males versus females is by the length of their antennae; males have longer antennae than females. C. maculatus are known to infest a range of legumes that are grown in the tropics and are a generalist in terms of what they infest. This makes it an effective model to study nutritional impacts as different storage legumes have very different nutritional content. We can therefore study the effects of presence or lack of protein thereof using natural legumes that vary largely in their pod shape and size.

Background

Observations are key to understanding behaviours and behavioural processes. Seed beetles are excellent model insects to study. They are also used in ecological and evolutionary biology research across many scientific research groups. Seed beetles are also pests of stored products. They infest a range of storage legumes and negatively impact global food security. Many developing countries produce (e.g. India) these legumes as food for human consumption and they also form a cheap source of protein as part of their diet. When seed beetles infest the stored legumes meant for human consumption they render them unfit for human consumption. Thus, understanding the behaviours of such serious food pests is warranted in order to come up with strategies to control them.

Many research labs use Bruchid beetles as a model system to study evolution and evolutionary processes. It is very popular among evolutionary biologists who study sexual conflict coevolution. Males, whose intromittent organ, is spiny and chitinous, can inflict damage to the female reproductive tract during copulation. Damage is visible when the female reproductive tracts are dissected and observed under a brightfield microscope.

Some research labs that use Bruchid beetles as a model system in their research are:

Paul Eady in Lincoln (UK): http://staff.lincoln.ac.uk/peady

Charles Fox’s lab in Kentucky (US): http://www.uky.edu/~cfox/

Goran Arnqvist (Uppsala): http://arnqvist.org/

Methodology

Using a laboratory based strain of C. maculatus, which was kindly given by a Dr N. Bakthavatsalam of the National Bureau of Agricultural Insect Resources (Bangalore, India), we carried out observations in groups of 2–4 to ascertain the common behaviours elicited by the pairs. Adults were acquired directly from the stock population and sexed before being introduced into the observation arena. We observed males and females in a cylindrical glass jar for approximately 45 minutes and noted down the behaviours. All behavioural observations were carried out under room temperature (~27°C to 28°C). Before the experimental observations began, we added 5 types of beans into the cylindrical glass jar and the five types of beans were red kidney bean, black-eyed beans, mung beans, rose cocoa beans and chick peas. Behaviours elicited during the 45-minute observation were recorded and how many time the beetles came into contact was also recorded.

Data shown here is group data pooled and the counts indicate the number of groups reporting these behaviours across the different foundations. Graphs were produced in “R” (R Core Team 2015) using “ggplot2” (Wickham and Chang 2015).

Results

The common behaviours observed during the 45-minute group observations were copulatory, oviposition (egg laying), preening and seed preference behaviours (which was either male/female choosing to remain on a particular seed). Across all these group observational studies, Foundation 5-Saturday (see Table 4), were the only group that reported same-sex behaviour in this strain of beetles. Flight and Mating (copulatory) behaviours were reported across all the foundation groups during their observations (see tables 1-5).

Table 1. Data collected by F2 (Sun)

Table 2. Data collected by F4 (Sat)

Table 3. Data collected by F5 (Sun)

Table 4. Data collected by F5 (Sat)

Table 5. Data collected by F7 (Sun)

Discussion

The strain of C. maculatus beetles used in this study showed a range of behaviours that are commonly observed and reported in scientific studies published in peer-reviewed research papers. Copulatory and flight behaviours were readily observed across all the foundation study groups as soon as the pairs were exposed to the experimental study environments. Many Bruchid beetles currently used for scientific studies are polyandrous wherein males will readily mate with several females within a short span of time. These behaviours observed and reported here are in line with such studies (Messina and Renwick 1985; Cope and Fox 2003).

One aspect of this study that we aim to improve for future sessions is to replicate the natural environments, where the beetles experience intense competition between males that frequently interrupt mating behaviour. This is known to impose intense selection pressures on males to optimize mating duration. We aim to do this by constructing environments that will take into account the presence and absence of intra-specific rivals within the experimental arena. This will hopefully enable us to observe and capture complex intra-specific fighting behaviour to obtain mates that are prevalent in many insect systems. We also aim to simulate different environmental conditions (e.g. varying temperatures) to observe how mating behaviour and with it other behaviours are affected by gradually increasing/decreasing their ambient temperature where they operate and capture the frequency of their behaviours elicited.

References

• Messina, F.J. and Renwick, J.A.A. 1985. Ability of ovipositing seed beetles to discriminate between seeds with differing egg loads. Ecological Entomology, 10(2), pp.225-230.

• Cope, J.M. and Fox, C.W. 2003. Oviposition decisions in the seed beetle, Callosobruchus maculatus (Coleoptera: Bruchidae): effects of seed size on superparasitism. Journal of Stored Products Research, 39(4), pp.355-365.

• R Core Team. 2015. R: a language and environment for statistical computing, Vienna, Austria.

• Wickham, H. and W. Chang. 2015. ggplot2: an implementation of the grammar of graphics. Available at https://cran.r-project.org/web/packages/ggplot2.